Folding protein molecules. Why the protein chain finds the only correct laying among many options. What is common in protein and universe
In the process of the synthesis of polypeptide chains, they are transported through membranes, when assembling oligomeric proteins, intermediate unstable conformations arise, prone to aggregation. On the newly synthesized polypeptide there are many hydrophobic radicals, which in the three-dimensional structure are hidden inside the molecule. Therefore, at the time of the formation of a native conformation, the reaction-capable amino acid residues of some proteins should be separated from the same groups of other proteins.
In all known organisms from prokaryotov to higher eukaryotes, proteins were found, capable of binding to proteins that are in an unstable, prone to aggregation. They are able to stabilize their conformation, providing folding proteins. These proteins were called "Chaperons".
1. Chaperons classifications (W)
In accordance with the molecular weight, all chaperons can be divided into 6 main groups:
high molecular weight, with a molecular weight of 100 to 110 kD;
W-90 - with molecular weight from 83 to 90 cd;
W-70 - with a molecular weight from 66 to 78 kD;
low molecular weight chaperons with molecular weight from 15 to 30 kD.
Among the chapers are distinguished: constitutional proteins (high basal synthesis of which does not depend on stressful effects on the cells of the body), and inducible, the synthesis of which is weak under normal conditions, but during stressful effects on the cell increases sharply. Inductable shaperons refer to "heat shock proteins", the rapid synthesis of which is noted in almost all cells that are subjected to any stressful effects. The name "heat shock proteins" arose as a result of the fact that for the first time these proteins were found in cells that were exposed to high temperature.
2. The role of chaperons in Folding proteins
In the synthesis of proteins, the N-terminal region of the polypeptide is synthesized earlier than the C-terminal region. To form a protein conformation, its complete amino acid sequence is needed. Therefore, during the synthesis of protein on the ribosome, the protection of reactive radicals (especially hydrophobic) is carried out with W-70.
W-70 is a high-circuit class of proteins, which is present in all cells of the cell: cytoplasm, core, er, mitochondria. In the region of the carboxyl end of the single polypeptide chain of the chapers, there is a plot formed by the amino acids in the form of a groove. It is able to interact with the sections of protein molecules and deployed polypeptide chains with a length of 7-9 amino acids enriched with hydrophobic radicals. In the synthesizing polypeptide chain, such sites are encountered around every 16 amino acids.
Folding of many high molecular weight proteins having a complex conformation (for example, a domain structure) is carried out in a special space formed by W-60. W-60 function in the form of an oligomeric complex consisting of 14 subunits (Fig. 1-23).
EC-60 form 2 rings, each of which consists of 7 subunits connected to each other. The S-60 subunit consists of 3 domains: apical (top), intermediate and equatorial. The top domain has a number of hydrophobic residues facing the rings formed by subunits. Equatorial domain has a plot of binding to ATP and has ATP-azna activity, i.e. It is capable of hydrolyzing ATP to ADP and H 3 PO 4.
The shaperone complex has a high affinity for proteins, on the surface of which there are elements characteristic of non-corned molecules (primarily areas enriched with hydrophobic radicals). Finding into the cavity of the chaperone complex, the protein binds to hydrophobic radicals of the apical sections of W-60. In a specific medium of this cavity, in isolation from other cell molecules there is a bust of the possible conformation of the protein, until the only one, the energy of the most profitable conformation is found.
The release of protein with the formed native conformation is accompanied by hydrolysis of ATP in the equatorial domain. If the protein did not acquire a native conformation, then it re-communicates with the shaperone complex. Such a shapernese-dependent folding of proteins requires the costs of a large amount of energy.
Thus, the synthesis and folding of proteins proceed with the participation of different groups of chapers that prevent the unwanted interactions of proteins with other cell molecules and accompany them to the final formation of the native structure.
4. Diseases associated with violation of folding proteins
Calculations have shown that only a small part of theoretically possible variants of polypeptide chains can take one stable spatial structure. Most of these proteins can take many conformations with approximately the same Gibbs energy, but with different properties. The primary structure of the majority of known proteins selected by evolution ensures the exceptional stability of one conformations.
However, some water soluble proteins with conditions change can acquire the conformation of poorly soluble capable of aggregation of molecules forming fibrillated sediments in the cells, referred to as the amyloid (from LAT. amylum -starch). Just like starch, amyloid deposits are detected when painting with iodine fabric. This can happen:
with hyperproduction of some proteins, resulting in their concentration in the cell;
if in their cells or the formation of proteins in them, capable of influence the conformation of other protein molecules;
when activating the proteolysis of normal proteins of the body, with the formation of insoluble, inclined to aggregation of fragments;
as a result of point mutations in the protein structure.
As a result of the deposition of amyloid in organs and tissues, the structure and function of cells are disturbed, their degenerative changes and the growth of connective tissue or glial cells are observed. Diseases are developing, called amyloiders. For each type of amyloidosis, a certain type of amyloid is characteristic. Currently, more than 15 such diseases are described.
Alzhemer's disease
Alzhemer's disease is the most frequently noted? -Amyloidosis of the nervous system, as a rule, striking people of old age and characterized by the progressive disorder of memory and full degradation of the individual. In the brain tissue is postponed? -Amylid - protein, forming insoluble fibrils, disturbing the structure and functions of nerve cells. ? -amilaid is a product of changes in the conforms of the normal protein of the human body. It is formed from the larger predecessor with partial proteolysis and is synthesized in many tissues. ? -Amilaid, in contrast to its normal predecessor containing a lot? -Spiral sections, has a secondary? -Calate structure, aggregates with the formation of insoluble fibrils, resistant to the action of proteolytic enzymes.
The reasons for the violation of the folding of native proteins in the brain tissue still have to be found out. Perhaps, with age, the synthesis of chaperons is reduced, capable of participating in the formation and maintenance of native protein conforms, or the activity of proteases increases, which leads to an increase in protein concentration, prone to change the conformation.
Prion diseases
Prions are a special class of proteins with infectious properties. Finding into the human body or spontaneously arising in it, they are able to cause severe incurable CNS diseases, called prion diseases. The name "Prions" comes from the abbreviation of the English phrase proteinaceous Infectious Particle- protein infectious particle.
The bottom protein is encoded by the same delented as its normal analogue, i.e. They have an identical primary structure. However, two proteins have a different conformation: the bottom protein is characterized by a high content? -Things, while normal protein has a lot? -Spiral plots. In addition, the booby protein is resistant to the action of proteases and, falling into the brain fabric or forging there spontaneously, contributes to the conversion of a normal protein into the subrink as a result of inter-brass interactions. The so-called "core of polymerization", consisting of aggregated prion proteins, to which new normal protein molecules are capable of joining. As a result, conformational rearrangements characteristic of prion proteins occur in their spatial structure.
There are cases of hereditary forms of prion diseases caused by mutations in the structure of this protein. However, it is possible to infect a person with prion proteins, resulting in a disease that leads to the death of the patient. Thus, Kuru is an union disease of the Aboriginal New Guinea, the epidemic character of which is associated with traditional cannibalism in these tribes and the transmission of infectious protein from one individual to the other. In connection with the change in the image of their life, this disease has practically disappeared.
A stunning game developed scientists from the University of Washington (USA). The program called Fold.It is a model for folding proteins in three-dimensional structures. Gamer must try to make it the most successful way. The program will be loaded with real data on these, just invented proteins, which are incomprehensible as folded. The results will go through the Internet into the processing center, where they will be checked on a supercomputer (it will be from autumn, and so far the program has already laid down riddles, so now it acts as a simulator).
In fact, all gamers of our world spend billions of human-hours for useless for humanity Games such as WOW, Counter-Strike or Solitaire "Kosyanka". At the same time, they could use intelligence more efficiently: for example, turning the proteins on the screen of their monitor. This is also interesting in its own way.
One of the developers of the game, Professor Biochemistry David Baker, sincerely believes that somewhere in the world there are talents who have a congenital ability to calculate 3D models of proteins in the mind. Some 12-year-old boy from Indonesia will see the game and will be able to solve the tasks that are not even a supercomputery. Who knows, maybe such people really have?
Each protein (in the human body there are more than 100,000 species) is a long molecule. To predict what an intricate form will take this molecule in certain conditions (and whether it is capable of incurred at all as a steady form) - the task of the highest degree of complexity. Computer modeling is a resource-sensitive process, but at the same time critical in pharmaceuticals. After all, not knowing the shape of the protein cannot simulate its properties. If these properties are useful, then proteins can be synthesized and on their base to make new effective preparations, for example, for the treatment of cancer or AIDS (the Nobel Prize is guaranteed in both cases).
Currently, hundreds of thousands of computers in a distributed computer network work on the compelling of the model of each new protein molecule, but scientists from the University of Washington offer another way: not a stupid bust of all options, but intellectual brainstorming through a computer game. The number of options is reduced by an order of magnitude, and the supercomputer will make the right parameters of the Folding much faster.
In three-dimensional "entertainment" fold.it can play all: even children and secretaries that have no idea about molecular biology. The developers tried to make such a game so that it was interesting to everyone. And the result of the game may well be the basis for the Nobel Prize and save the life of thousands of people.
The program is released in versions under Win and Mac. 53 MB Distribution Distribution
Biological chemistry LELEVICH Vladimir Valerianovich
Folding
Folding proteins - the process of folding the polypeptide chain into the correct spatial structure. In this case, there is a convergence of remote amino acid residues of the polypeptide chain, leading to the formation of a native structure. This structure has unique biological activity. Therefore, Folding is an important stage of transformation of genetic information into the mechanisms of cell functioning.
Structure and functional role of chaperons in Folding proteins
In the process of the synthesis of polypeptide chains, they are transported through membranes, when assembling oligomeric proteins, intermediate unstable conformations arise, prone to aggregation. On the newly synthesized polypeptide there are many hydrophobic radicals, which in the three-dimensional structure are hidden inside the molecule. Therefore, at the time of the formation of a native conformation, reactive amino acid residues of some proteins should be separated from the same groups of other proteins.
In all known organisms from prokaryotov to higher eukaryotes, proteins were found, capable of binding to proteins that are in an unstable, prone to aggregation. They are able to stabilize their conformation, providing folding proteins. These proteins were called Shaperonov.
Chaperon Classification (W)
In accordance with the molecular weight, all chaperons can be divided into 6 main groups:
1. High molecular weight, with a molecular weight from 100 to 110 kDa;
2. WC-90 - with molecular weight from 83 to 90 kDa;
3. W-70 - with a molecular weight from 66 to 78 kDa;
6. Low molecular weight chaperons with a molecular weight from 15 to 30 kDa.
Among the chapers are distinguished: constitutional proteins (high basal synthesis of which does not depend on stressful effects on the cells of the body), and inducible, the synthesis of which is weak under normal conditions, but during stressful effects on the cell increases sharply. Inductable shaperons belong to the "heat shock proteins", the rapid synthesis of which is noted in almost all cells that are subjected to any stressful effects. The name "heat shock proteins" arose as a result of the fact that for the first time these proteins were found in cells that were exposed to high temperature.
The role of chaperons in Folding proteins
In the synthesis of proteins, the N-terminal region of the polypeptide is synthesized earlier than the C-terminal region. To form a protein conformation, its complete amino acid sequence is needed. Therefore, during the synthesis of protein on the ribosome, the protection of reactive radicals (especially hydrophobic) is carried out with W-70.
WCH-70 is a high-circuit class of proteins that is present in all sections of the cell: cytoplasm, core, mitochondria.
Folding of many high molecular weight proteins having a complex conformation (for example, a domain structure) is carried out in a special space formed by W-60. W-60 function in the form of an oligomeric complex consisting of 14 subunits.
The shaperone complex has a high affinity for proteins, on the surface of which there are elements characteristic of non-corned molecules (primarily areas enriched with hydrophobic radicals). Finding into the cavity of the chaperone complex, the protein binds to hydrophobic radicals of the apical sections of W-60. In a specific environment of this cavity, in isolation from other cell molecules, the choice of possible protein conformations occurs until the only one, the energetically most favorable conformation is found.
The release of protein with the formed native conformation is accompanied by hydrolysis of ATP in the equatorial domain. If the protein did not acquire a native conformation, then it re-communicates with the shaperone complex. Such a shaper-dependent folding of proteins requires the costs of a larger amount of energy.
Thus, the synthesis and folding of proteins proceeds with the participation of different groups of chapers that prevent the unwanted interactions of proteins with other cell molecules and accompany them to the final formation of the native structure.
The role of chapers in the protection of cell proteins from denaturing stressful effects
Chaperons involved in the protection of cellular proteins from denaturing effects, as mentioned above, refer to the proteins of heat shock (BTSH) and in the literature are often denoted as HSP (HEAT SHOCK Protein).
Under the action of various stress factors (high temperature, hypoxia, infection, UFO, the change in the middle of the medium, the change in the molarity of the medium, the effect of toxic chemicals, heavy metals, etc.) in the cells, the synthesis of BTSH is enhanced. Having a high affinity for hydrophobic sections of partially denatured proteins, they may interfere with their complete denaturation and restore the native conformation of proteins.
It has been established that short-term stress exposure increases the production of BTSH and increase the resistance of the body to long-term stressful effects. Thus, short-term ischemia of the heart muscle during the run period with moderate training significantly increases myocardial stability to long-term ischemia. Currently, the search for pharmacological and molecular biological methods for activating the synthesis of BTSH in cells is considered promising research in medicine.
Diseases associated with violation of folding proteins
Calculations have shown that only a small part of theoretically possible variants of polypeptide chains can take one stable spatial structure. Most of these proteins can take many conformations with approximately the same Gibbs energy, but with different properties. The primary structure of the majority of known proteins selected by evolution ensures the exceptional stability of one conformation.
However, some water-soluble proteins with conditions change can acquire the conformation of poorly soluble, capable of aggregation of molecules forming fibrillar deposits in the cells, referred to as amyloid (from Lat. Amylum - starch). Just like starch, amyloid deposits are detected when painting with iodine fabric.
This can happen:
1. In case of hyperproduction of some proteins, resulting in their concentration in the cell;
2. When in cells or the formation of proteins in them, capable of influence the conformation of other protein molecules;
3. When activating the proteolysis of normal proteins of the body, with the formation of insoluble, inclined to aggregation of fragments;
4. As a result of point mutations in the protein structure.
As a result of the deposition of amyloid in organs and tissues, the structure and function of cells are disturbed, their degenerative changes and the growth of connective tissue cells are observed. Diseases are developing, called amyloidosis. For each type of amyloidosis, a certain type of amyloid is characteristic. Currently, more than 15 such diseases are described.
Article for Competition "Bio / Mol / Text": Proteins are the main biological molecules. They perform a variety of diverse functions: catalytic, structural, transport, receptor and many others. Even the well-known DNA plays only the role of "flash drives", keeping information about proteins, while proteins themselves "files". Life on Earth can be called protein. But do we really know about the structure and functioning of these substances? Until now, the secret remains filding protein - the process of spatial packaging of a protein molecule, the adoption by a protein of a strictly defined form in which it performs its functions.
The general sponsor of the competition, according to our crowdfund, became an entrepreneur Konstantin Sinyushin why he has a huge human respect!
The sponsor of the prize of spectator sympathies was the firm "Atlas".
Sponsor of publishing this article - Lev Makarov.
Proteins - biopolymers, which can be compared with beads, where the beads are amino acids, interconnected by peptide bonds (hence another name of proteins - polypeptides). In the cell, proteins are synthesized on special molecular machines - ribosomes. Leaving the ribosomes, the polypeptide chain is folded, and the protein takes a certain conformation, that is, a spatial structure (Fig. 1). It is vital that the protein is present in the body in a certain form, that is, the conformation must be "proper" (native). The process of folding the protein and is called the Folding (from the English. folding. - folding, laying; Note that the term "Folding" is applicable not only to proteins). The most interesting thing is that information about the three-dimensional structure is "laid" in the very sequence of amino acids. Thus, the protein to take a native structure is only required to know, in which sequence and what amino acid residues are present in it. For the first time, this was proved in 1961 by Christian Anfinsen on the example of bovine pancreatic ribonuclease (Fig. 2). It should be said that, in addition to proteins, whose spatial structure is strictly determined by the amino acid sequence, there are so-called unstructured proteins ( intrinsically Unfolded Proteins, IDPS): Some fragments of such molecules, and sometimes whole molecules, are able to receive many possible conformations at once, and they are all energy "equivalent", and such proteins are quite often found in nature and perform important functions. There is another type of folding, occurring with the help of special proteins - chaperons, but a little later.
Figure 1. Handling folding of a small α-spiral domain. The collapse of the polypeptide chain of many proteins begins in the ribosome during the transmission of the protein (that is, its synthesis). The ripening protein comes out of the ribosomes through a special tunnel (in the figure - a darkened area in a large subunit), which is an important factor in the collapse of the chain, and the C-end of the chain (containing a carboxyl group) is fixed in the ribosome, and n-end (containing an amino group) "is moving "To the exit and" hangs "from it, when 30-40 amino acid residues accumulate in the tunnel. In the tunnel, compactized immature structures, α-helix, β-studs and small α-spiral domains can be formed. Cutment Folding passes in two stages: at first the incomplete chain ( U, unfolded) goes into a compactized state ( C, Compacted), which then acquires a native structure ( N, Native.).
Figure 2. Bullway pancreatic ribonucleases and scientists who studied it. but - Bull Pancreatic Ribonuclease. For the study of the structure of this enzyme Anfinsen ( Anfinsen.) (b. ), Moore ( Moore.) (in ) and Stein (Stein.) (g. ) Received the Nobel Prize in Chemistry (1972) ,. On the example of this protein, the phenomenon of refolding was shown for the first time - the spontaneous formation of the tertiary structure after denaturation (that is, destruction). The value of protein folding is that it leads to the formation of a strictly defined (native) protein structure in which it functions. For example, in the experience of the Anfinsen ribonuclease as a result of refolding, it restored its enzymatic activity, that is, it began to catalyze the biochemical reaction again. In order for this enzyme to work, five amino acid residues should be gathered in a single catalytic center (one piece of space) from completely different places: histidine (12), lysine (41), threonine (47), histidine (119) and phenylalanine ( 120).
model from the PDB database (PDB ID 5D6U), portraits of scientists from the site ru.wikipedia.org
The relevance of the problem
The problem is that humanity with all its computational capacities and the arsenal of experimental data has not yet learned to build models that would describe the protein folding process and predict the three-dimensional protein structure based on its primary structure (that is, the amino acid sequence). Thus, there is still no complete understanding of this physical process.
The explosive growth of genomic projects led to the fact that more and more genomes are sequenced, and the corresponding DNA sequences and RNAs fill the databases on the exponential. In fig. 3 shows an increase in the number of amino acid sequences, as well as an increase in the number of well-known protein structures in the period from 1996 to 2007. It is clearly seen that the number of well-known structures is significantly less than the number of sequences. At the time of writing this article (August 2016), the number of sequences in the UniParc database is more than 124 million, while the number of structures in the PDB database ( Protein Data Bank.) - Only a little more than 121 thousand, which is less than 0.1% of all known sequences, and the gap between these two indicators is rapidly growing and will likely grow further. Such a strong lag is associated with the relative complexity of modern methods for determining structures. At the same time know them is very important. Therefore, the question of the use of computing methods to predict protein structures by their sequences is now sharp. In 2005, an authoritative magazine Science Recognized the problem of folding protein one of the 125 largest problems of modern science.
Figure 3. Comparison of the growth rates of the number of well-known sequences and structures from 1996 to 2007. On the horizontal axis, years are indicated on the left vertical - the number of sequences in millions ( solid line), on the right vertical - the number of structures in millions ( dotted line). Clearly seen the backlog of the number of known structures on the number of sequences. To date, the gap grew even stronger.
After reading the human genome, many human genes became known and, consequently, the amino acid sequences encoded by them. However, this does not mean that we know the functions of all genes, in other words, we do not know the function of proteins encoded by these genes. It is known that in many respects the proteins function can be predicted by their structure, although not always ,. Therefore, the cherished dream is the ability to predict the structure and, as a result, the protein function along the nucleotide sequence of the gene.
What is done to solve the problem?
Incorrectly, however, think that we do not know anything at all. Of course, a large number of facts about the Folding have been accumulated, the patterns of this process are known, various methods of its modeling have been developed. To keep track of success achieved on the way to solving the Folding problem, an international competition for predicting the spatial structure of protein molecules - Casp was created ( Critical Asesement of Techniques for Protein Structure Prediction), goes every two years (now the competition is currently twelve, it began in April and will end in December 2016). In this contest, the researchers compete, who will better predict the structure of the protein on its amino acid sequence, and the competition passes using a double-blind method (at the time of the competition, the structure of the "riddle" is simply unknown; its definition is completed every time at the end of the competition). So far, the structures of target proteins were not exactly predicted.
There are two groups of structural prediction methods.
TO first These are the so-called methods of modeling "from scratch" (aB Initio, de Novo, There are other synonymic terms) when the models are constructed only on the basis of the primary structure, without the use of comparative methods with already known structures, but using the entire accumulated understanding of biopolymers folding physics. The fundamental importance of these methods is that they help to understand the physicochemical principles of protein filding, to answer this burning question - why the protein turns out so, and not otherwise? However, the disadvantages of these methods are a very greater complexity of calculation and low accuracy. These methods require simplifications and approximations, and are also ineffective to predict the structures of large proteins. In 2007, due to modeling methods de Novo. For the first time with high accuracy, the structure of one of the bacteria proteins was determined Bacillus Halodurans.but this protein is relatively small (112 amino acid residues), and for obtaining an accurate model, the capacity of more than 70,000 personal computers and a supercomputer was required; In addition, from the 26 models obtained, only one turned out to be accurate. Molecular dynamics methods (MD) allow you to describe molecular events and are able to trace the process of folding the protein into a native structure: in 2010, for the first time, it was possible to do this at the expense of the computing power of a specially created supercomputer Anton. .
Ko second Group of methods are related methods of comparable modeling. They are based on the phenomenon homology, that is, the generality of the origin of objects (organs, molecules, etc.). Thus, the "predictor" has the opportunity to compare the protein sequence, the structure of which must be modeled, with the template, that is, the protein, the structure of which is known and which is presumably a homologue, and on the basis of their similarity to build a model with subsequent adjustments (similar sequences are collapsed to the similar structures). These methods are now more popular, since the prediction of the structure of proteins is an important practical task, and to date, computational means appeared, databases, and it also became known that the number of possible options for stacking protein structures is limited, (Fig. 4). And let these methods do not remove the problem of protein folding, they are able to help solve specific practical tasks, while others are fighting over the study of more fundamental issues.
Figure 4. Dynamics of identifying new types of fildal (packaging options). At the horizontal axis, the time (years) is postponed, on the left vertical axis - the proportion of new fildards (in more detail on the tab) ( solid line), and on the right vertical axis - the total number of structures ( dotted line), classified in the CATH database. Note that this database is engaged in the structural classification of proteins, so it is fundamentally to know the possible types of protein fildren. It is clearly seen that over time more and more proteins are classified, but the number of Folding variants decreases.
It should be emphasized that modern methods of predicting protein structures require high computing power and are often carried out on supercomputers or using distributed computing networks, such as, for example, [Email Protected] and [Email Protected] . Everyone will be invited to participate in these projects: you only need to run the program on your computer until it is needed by the user.
Some regular fonding protein
Known some patterns of protein folding. Now it is believed that this process occurs in stages: first the linear chain having zero entropy is rapidly collapsing with education statistical Cluster - Entropic Folding . Then occurs hydrophobic collapse: Hydrophobic amino acid residues "hide" deep into the molecule, and hydrophilic - "settle" on the surface (see below). The result of this stage is the formation molten globule. After that, the formation of specific connections (see below), and the protein goes into the state of true globulaAt the same time free energy drops sharply.
The last stage does not occur during the Folding of unstructured proteins - Idps..
It should be noted that for each amino acid sequence, it is theoretically to assume many paths that it can go to achieve native conformation. However, it is known that the protein does not go through all possible options, but moves one of the possible paths defined for each sequence. If the protein tried all possible options, while the path from the simple sequence to the native state would exceed the existence time of the Universe (Levintal Paradox)! Of course, this does not occur: the taking time of the protein of the native structure is a fraction of a second. It looks like an assembly of Rubik's cube: From the state of the insufficient cube to the state of the assembled, you can come with many different ways, but the one who makes it faster and more efficient, that is, chooses a certain path, is defeated at the Cube Speed \u200b\u200bSpeed \u200b\u200bCompetitions. Actually find such a way - and there is the main task of modeling methods aB Initio. (see above). The answer to the fundamental question of the Folding will be not easy in the ability to unmistakably simulate structures, but, first of all, to know and substantiate the way to achieve a protein of a native state.
The value of the cotton Folding should be emphasized (Fig. 1), which was mentioned above, in the formation of the structure of the protein. Note that the presence of ribosomes on which protein is synthesized, imposes serious adjustments to the process of folding the chain. It should always be borne in mind when modeling the folding of natural proteins in vivo.. The channel, which turns out to be a growing chain, limits its conformational variability, and therefore far from all types of structures can be formed in it ,. In addition, the growing chain is constantly pushing forward (one amino acid residue with each act of translocation translocation, i.e., the formation of a new peptide bond and subsequent ribosome promotion), and therefore it will be logical to assume that the conformation of the chain in the ribosomous channel has such qualities as hardness and the vector, which corresponds to the properties of the α-helix. In addition, the mutual orientation of amino acid residues in two centers inside the ribosomes is always the same type (equivalent), which does not depend on the nature of these residues, which also seems to contribute to the formation of α-helix. Indeed, the α-helix is \u200b\u200bthe most typical element of the secondary structure of proteins. They were opened by Linus Pauling ( Liunus Pauling) and Robert Corey ( Robert Corey.) who, together with Walter Koltun ( Walter Koltun.) offered a new type of models of molecules.
At the same time, when the N-end (containing an amino group) of a growing protein chain comes out of the tunnel and immersed in the solution, the physico-chemical conditions of this medium begins to operate, and the protein begins to obey their rules.
The famous molecular biologist Academician Alexander Spirin in this regard marks three differences between the Folding in vitro. and in vivo.:
- First, the starting conformation is different: if at the experimental conditions, renaultation begins with a certain state of the deployed chain in the solution, then in the case of ribosome filding, it starts with some particular conformation provided by the ribosomal channel.
- Secondly, with the cotton Folding, the folding begins with the N-end, that is, the process of the Folding direction is directed, and in the case of the Folding without the participation of the ribosomes, the search for conformations is carried out at once the entire molecule.
- The third difference lies in the fact that in the case of the cotton folding, the C-end of the protein chain is fixed with a ribosome, relative to a large particle, which leads to stabilization of intermediate structures (see above), and in the case of refolding in vitro. This stabilization does not occur.
These considerations prove once again that biological issues cannot be solved "dry" through the use of bioinformatics methods. Even the most, it would seem, the verified computer models may be inaccurate if they are built without taking into account the factors actively operating in nature.
To solve the problem of the Folding, the so-called empirical potentials are developed: paired reasons of residues, hydrogen bonds, torsion corners, centers of side chains and many others ,. For example, the potential of solvation allows you to predict, inside or outside the protein will be an amino acid residue (respectively, bellulated or exposed) depending on its hydrophobicity. It is known that alone amino acids "love" water ( hydrophilic), they will most likely be located on the surface of the protein molecule, while others are "not like" ( hydrophobic) and "hide" into the more inaccessible for the solvent region of the molecule, leaving other residues (Fig. 5). The hydrophobic effect is of great importance in the folding of the protein.
Figure 5. The hydrophobicity of amino acids affects their spatial distribution (on the example of one of the human dehydrogenases). Hydrophilic amino acids are shown blue blossom, hydrophobic - red. It can be seen that the hydrophilic residues tend to be located on the solvent open for the solvent, while hydrophobic - in the closed areas of the molecule.
pDB database (PDB ID 5ICS)
An important aspect of the formation of a protein structure at all stages is the formation of links between radicals (lateral chains) of amino acid residues. They are different: hydrophobic, electrostatic and others. An interesting embodiment is the formation of disulfide bonds ("bridges") due to the interaction of cysteine \u200b\u200bside chains of sulfur chains. For example, in the famous ribonuclease, for the study of the structure of which the Nobel Prize was given, four such links. However, everything is not so simple here. If the protein chain includes two sulfur atoms belonging to cysteine, it is easy to say that one disulfide bridge may form. But if the sulfur atoms, for example, ten and, accordingly, five SS-links are formed, then we cannot definitely say which sulfur atoms will interact in pairs with each other (and the protein may). According to the calculations of Thomas Creiton ( Thomas Creighton) If in protein 5 disulfide bonds, the number of possible combinations is already 945, if such bonds 10, the number of options is 654,729,075, and with 25 disulfide bonds, this number exceeds 5 quadrillion quadrillion (more than 5.8 × 10 30). And only one option is implemented in the protein, and moreover is always the same! It should be noted that it is true for the self-organization of proteins in vitro. ("In a test tube", "in glass", that is, in the conditions of the experiment, and not in a living organism) in suitable conditions, and in vivo. (in the living organism) self-organization of disulfide bonds does not occur. Their education catalyzes a special enzyme - proteindisulfidisomeraza , or PDIwhich is also able to "correct" errors in case of improper formation of SS communication, thus adjusting the process of the Folding ,.
It is important to understand that the process of forming the final structure of the protein is not only in the simple folding chain. In cells, proteins are subjected to acetylation, glycosylation and many other modifications. Therefore, for example, the number of different amino acids in proteins exceeds the known 20 ("magical twenty", according to the articulated expression of the Nobel laureate of Francis Creek). In addition, for the formation of complex (oligomeric) proteins, the formation of specific bonds between individual proterars (for example, in the hemoglobin molecule, four protsometers, that is, separately synthesized chains). For many proteins, especially enzymes, the attachment of the prosthetic group is important, that is, a non-peculiar component. Other conversions may occur.
Many other patterns of protein folding are known. The veil is gradually lifted. However, the picture is still far from the holistic. The successes of the prediction of structures are only episodic. In this regard, the scientific community made the following curious step: it attracted a wide public to solving the issue, creating a game Foldit. . Anyone can take part in the global competition. The essence of the game is to minimize the protein chain as compact as possible, that is, to bring a protein molecule into such a state in which the free space inside the glider as little as possible - it is precisely in this form that proteins are present in nature (Fig. 6). From the point of view of thermodynamics, this state corresponds to a minimum of free energy ,. The more compact molecule is obtained than the less cavities and open hydrophobic areas, the more open hydrophilic areas, hydrogen bonds in the structures of the β-sheets, the less "collisions" of atoms, the greater the number of points the player is accrued. Thus, the largest number of points get a model with the smallest free energy. Most players Foldit. They have only small biochemical training either do not have it at all. The game is based on Rosetta algorithms and is not modeling structures de Novo.which, as the authors are correctly noticed, is still an extremely difficult problem.
Figure 6. Comparison of different forms of representation of models of protein structures (on the example of one of the human transfers). but - Form, clearly demonstrating the types of secondary structures. b. - a form showing the real location of the atoms of the protein molecule in space ( Space Fill). It is clearly seen that protein molecules are very compactized, there is little free space between atoms.
pDB database (PDB ID 5CU6)
Group of players Foldit. Takes part in Casp. The game has already shown its effectiveness in predicting structures and even greater efficiency in comparison with other methods, and also solved the serious scientific problem of the structure of the virus protease of immunodeficiency monkeys, which science could not solve for more than decades.
Speaking about the use of different methods and means to solve the problem under discussion, it should always be remembered that not all sequences can be rolled out strictly in a certain way. Probably, we, looking at the results to which the evolution came to date, we see only those sequences that can be folded, as they performed their functions well and were supported by the selection.
"Governess" for proteins - Chaperons
Speaking about Folding, we focused on the relative autonomy of this process: the protein molecule takes a certain conformation on the basis of its primary structure, and it occurs in specific (which is important) physicochemical conditions (acidity, temperature, solvent nature, etc.). Nevertheless, there should be no impression that the Folding is absolutely independent, especially for large proteins. We just mentioned the PDI enzyme that helps the squirrel to turn correctly. In addition to this enzyme, there are others (for example, PPI - peptidyl shed-cis / trans isomerase ). But enzymes is not the only group of proteins that helps to correctly turn into other proteins. There is another special group of proteins playing an important role in the Folding. They are called chapers.
Chaperons - complex proteins with conservative (that is, an evolutionary little changeable) mechanism of action, found in all kingdoms of wildlife. This is understandable: their role in the cell life is enormous. As mentioned above, the ripening protein chain comes out of the ribosome. She is still immature, and abides in the so-called "melted" state. Such immature molecules are subject to the evil influence of the environment: they can interact with other cellular proteins, forming the aggregates, which can lead to diseases, for example, Alzheimer's disease or Parkinson. But there is also a "right" course, which may (and should) be directed by the development of protein, is the path that will lead the molten globule into a native state. Here and help the chaperons, "podkarayuyu" and exciting protein chains at the exit of the ribosomal tunnel and thus guiding unripe proteins that are on a fateful crossroads in the right track. Chaperons are called so no accident: before in England, the an elderly experienced lady called that, which accompanied the young girl who first published under her leadership, and kept her from ill-conceived contacts. (The term "Shaperone" and is now used in close values.) Chaperons are not specific to different amino acid sequences of nascent chains, but they may distinguish mature proteins from immature and act on the latter.
The most important group of Chaperons - shaperonins. Their structure is interesting: they are kegs made up of two rings. The folding protein falls inside the shaperonin, and the "entrance" is closed by a special "cap" or the closure of the edges of the blocks from which the ring is consisting so that the protein molecule does not leave the chaperonin ahead of time (Fig. 7). In such a protected state, the protein can finally take a native conformation. So far, the processes occurring inside the chaperonin barrels are small.
Figure 7. Schematic representation of two types of shaperonins - I and II. but - type I Chaperonins are characteristic of bacteria (Chaperone Groel. has a barrel structure made up of two rings, in each - 7 "blocks"; inside the chaperonin - a chamber in which the conversion of the molten globule into the native; Barrel closes "lid" - Groes.); b. - Type II Chaperonins characteristic of archaees and eukaryotes (here each of the two rings consists of 8 "blocks"; the closure of the chamber does not occur due to the connection of the "cover", but according to the mechanism of the camera's lens).
It must be said that the chaperons not only participate in the folding of ripening chains, but also help "broken" protein structures that occurred in a cell as a result of certain impacts, again take the correct conformation. The most typical reason for such "breakdowns" - thermal shock, that is, raising the temperature. In this regard, other chapels are often used - heat shock proteins ( hEAT SHOCK PROTEINS, HSP) or stress proteins. Shaperons perform other important functions in a cell, for example, protein transport through membranes and an assembly of oligomeric proteins.
Conclusion
So, the following conditions are strictly needed for the folding of the protein: primary structure, specific physicochemical conditions, as well as two groups of auxiliary proteins - specific enzymes and nonspecable shaperons.
Summarizing, let's say that protein filding is one of the central problems of modern biophysics. And although there is a large arsenal of data on this phenomenon, it is still uninterrupted, which is ultimately expressed, ultimately, in the impossibility of predicting the three-dimensional structure based on the amino acid sequence (this also applies to large, including oligomeric, proteins). Successes in this area, and especially modeling de novo. (2005). Science. 309
, 78–102;
Folding is the process of laying an elongated polypeptide chain into the correct three-dimensional spatial structure. To ensure the Folding, a group of auxiliary proteins called Shaperons (Chaperon, Franz is used. - Satellite, Nannik). They prevent the interaction of newly seated proteins with each other, isolate the hydrophobic sections of proteins from the cytoplasm and "remove" them inside the molecule, the protein domains are properly positioned. Chaperons are represented by families consisting of homologous structures and proteins functions that differ in the nature of the expression and the presence in different cell compartments.
In general, the chaperons contribute to the transition of the structure of proteins from the primary level to the tertiary and quaternary, but they are not part of the final protein structure.
Novosynthesed proteins after accessing with ribosomes for proper functioning should be laid into stable three-dimensional structures and remain such throughout the entire functional life of the cell. Maintaining the quality control of the structure of the protein and is carried out by chapers catalyzing the laying of polypeptides. The assembly of polyproteins and laying of multi-cell complexes is also carried out by chapers. Shaperons bind to hydrophobic areas of incorrectly laid proteins, help them to curl and achieve a stable native structure and, thus, prevent their inclusion in insoluble and non-functional aggregates. During its functional life, protein may be subjected to various stress and denaturation. Such partly denatured proteins may be, firstly, the target of proteases, secondly, aggregate and, thirdly, to fit into the native structure with the help of chapers. The balance and effectiveness with which these three processes occur are determined by the ratio of components involved in these reactions.
Transport of many proteins from one compartment to another.
Participation in signaling paths. For example, the presence of HSP70 is necessary to activate phosphatase, which by dephosphorylation inhibits the jnk proteincinase, the component of the stress-induced apoptosis signal, i.e. HSP70 is part of the antipoptotic signaling path.
Regulation of functions of various molecules. For example, the steroid receptor located in the cytoplasm is associated with HSP90; The ligand falling into the cytoplasm joins the receptor and displaces the chaperone from the complex. After that, the ligand receptor complex acquires the ability to bind to DNA, migrates to the kernel and performs the transcription factor function.
In disruption of the functions of the chaperons and the absence of folding in the cell, protein sediments are formed - amyloidosis develops. The amyloid is a glycoprotein, the main component of which is fibrillar proteins. They form fibrils that have a characteristic smulosicroscopic structure. Fibrillar amyloid proteins are heterogeneous. There are about 15 variants of amyloidosis.
Prions
It seems that Folding with the participation of Foldas and Chaperon leads to the correct one. The most optimal structure in the energy and functional relationships. However, it is not. There is a group of severe neurological diseases due to improper folding of one, quite a certain protein.
It is known that PRP can exist in two conformations - "Healthy" - PRPC, which it has in normal cells (C - from the English. Cellular - "cellular"), in which the alpha spirals prevail, and "pathological" - PRPSC, actually Prion (SC-from SCRAPIE), for which the presence of a large number of beta-heavyness is characterized.
The arrival protein with an anomalous three-dimensional structure is capable of directly catalyze the structural transformation of the homologous normal cellular protein in itself a similar (arrivals), joining the target protein and changing it conformation. As a rule, the prion state of the protein is characterized by the transition of the α-helix of the protein in the β-layers.
If you get into a healthy cell, PRPSC catalyzes the transition of the Cell PRPC to the subront conformation. The accumulation of the prion protein is accompanied by its aggregation, the formation of highly ordered fibrils (amyloids), which in the end leads to the death of the cell. The released prion seems to be able to penetrate into neighboring cells, also causing their death.
PRPC protein functions in a healthy cell - maintaining the quality of myelin shell, which in the absence of this protein gradually thinned. In the norm, PrPC protein is associated with a cell membrane, glycosylated by the residue of sialic acid. It can perform cyclic transitions inside the cell and back to the surface during endo and exocytosis.
Until end, the mechanism of spontaneous occurrence of prion infections is not clear. It is considered (but not yet completely proven) that the prions are formed as a result of errors in the biosynthesis of proteins. Mutations of genes encoding the arrivals (PRP), broadcast errors, proteolysis processes are considered to be the main candidates for the mechanism of the occurrence of prions.
Thus, the prions are a special class of infectious agents, purely protein, non-nucleic acids that cause severe diseases of the central nervous system in humans and a number of higher animals (t. N. "Slow infections").
There is data that gives reason to believe that the prions are not only infectious agents, but also have functions in normal bioprocesses. For example, there is a hypothesis that through the prions is carried out by the mechanism of genetically determined stochastic aging.
Prions - the only known infectious agents whose reproduction occurs without the participation of nucleic acids.
In the second half of the 20th century, doctors faced an unusual human disease - gradually the progressive destruction of the brain, resulting from the death of nerve cells. This disease got the name of the spongy encephalopathy. Similar symptoms were known for a long time, but they were not observed in a person, but in animals (scraping sheep), and for a long time they did not find a sufficient reasonable connection between them.
A new interest in studying them arose in 1996, when a new form of the disease appeared in the UK, denoted as "a new version of Creitzfeldt-Jacob's disease.
An important event was the spread of "cow's rabies" in the UK, the epidemic of which was first in 1992-1993, and then in 2001 covered several European states, but nevertheless, the export of meat to many countries was not discontinued. The disease is associated with the use of "obvious" bone flour in feed and premixes made from the carcass of the fallen or sick animals, possibly and did not have obvious signs of the disease.
Ways to transfer the causal factor of the disease, the mechanisms of penetration of prions into the body and the pathogenesis of the disease are not yet sufficiently.
Mammal Prions - Sovipitals of Spongean Encephalopathy
Scraping Sheep and Goat Scrolling OvPrPSC
Transmissive encephalomyopathy mink (TEN) Prion TEN and MKPRPSC
Chronic Wasting Disease (CWD) Deer and Moose CWD Prick MDEPRPSC
Spongeous encephalopathy cattle (HECC) cow prion bovprpsc
Spongy Encephalopathy Feline (HEK) Cats Prion GEK FEPRPSC
Spongeous Encephalopathy Exotic Unit (EUE) Antelope and Big Kund EUE Prick NyaprPSC
Kuru People Prick HuprPSC
Crazzfeld-Jacob's disease (BKY) People Prick BKA HUPRPSC
(NEW) Variant Creutzfeldt-Jakob Disease (VCJD, NVCJD) People VCJD HUPRPSC
Herstrene-Strocera-Sheinkers syndrome (GSS) GSS people arrive HuprPSC
Fatal Family Insomnia (FSB) People Prick FSB HUPRPS
A person can become infected with the prions contained in food, as they are not destroyed by the enzymes of the digestive tract. Since they are not adsorbed by the intestinal walls, they can penetrate the blood only through damaged tissues. Ultimately, they fall into the central nervous system. So the new version of Creitzfeldt-Jacob (NVCJD) disease (NVCJD) is transferred, which people are infected after eating beef containing nervous fabrics from cattle heads, sick bullish encephalopathy (BSE, cow's rabies).
In practice, the possibility of prions infect the body with air-droplet mice is proved.
Prions can penetrate the body and parenteral. Cases of infection with intramuscular administration of drugs made of human pituitary gland (mainly hormones of growth for the treatment of dwarfs), as well as brain infection with tools for neurosurgical operations, since prions are resistant to currently used thermal and chemical sterilization methods. This shape of Creitzfeldt-Jacob's disease is indicated as non-hydrogen (1cjd).
With certain, unknown conditions, a spontaneous transformation of the prion protein can occur in the human body in the prion. So the so-called Cratetzfeldt-Jacob (SCJD) disease occurs, first described in 1920, independently from each other by Gansa Gerhard Kreitzfeldt and Alphonace Maria Jacob. It is assumed that the spontaneous occurrence of this disease is associated with the fact that in the normal body in the human body there is a small number of prions, which are effectively eliminated by the Cellular apparatus of Golgi. The violation of this ability of "self-cleaning" cells can lead to an increase in the level of prions above the permissible boundary of the norm and to their further uncontrollable distribution. The reason for the occurrence of sporadic disease Creitzfeldt-Jacob according to this theory is a violation of the function of the Golgi's device in cells.
A special group of prion diseases are hereditary (congenital) diseases caused by a mutation of a prion protein gene, which makes an emerging prion protein more susceptible to spontaneous change in the spatial configuration and turn them into prions. The hereditary form of the hereditary diseases includes the hereditary form of Craitzfeldt-Jacob (FCJD), which is observed in a number of countries in the world. With prion pathology, the highest concentration of prions was found in the nervous tissue of infected people. Prions are found in lymphatic tissue. The presence of prions in biological fluids, including saliva, has not yet been uniquely confirmed. If the idea of \u200b\u200bthe constant emergence of a small number of prions is true, then it can be assumed that new, more sensitive diagnostic methods will open this number of prions scattered over various tissues. In this case, however, it will be discussed about the "physiological" level of prions, which are not a threat to humans.
