Proteins

Mystery of our body

Although it is possible to deduce the primary structure of a protein from a gene's sequence, its tertiary structure cannot be determined (although it should become possible to make predictions when more tertiary sequences are submitted to databases). It can only be determined by complex experimental analyses and, at present, this information is only known for about 10% of proteins


By MARIJA MITROVIĆ
from Belgrade, SERBIA


Name of the protein, from Greek proteois which means the first, signify its importance in our organism.. Proteins are fundamental components of all living cells: our own, the bacteria that infect us, the plants and animals we eat. The hemoglobin that carries oxygen to our tissues, the insulin that signals our bodies to store excess sugar, the antibodies that fight infection, the actin and myosin that allow our muscles to contract, and the collagen that makes up our tendons and ligaments (and even much of our bones)-all are proteins maid in by our organism. Beside that protein based antibiotics and vaccines help us to fight disease, and we warm and protect our bodies with clothing and shoes that are often protein in nature (e.g. wool, silk and leather). Although, they are generally regarded as beneficial, proteins can also be one of the strongest poisons existing in nature, e.g. venoms of many snakes, tetanus or Botulinum toxin A. A teaspoon of this toxin, very popular in aesthetic surgery, would be sufficient to kill a fifth of the world's population.

To make proteins, 'machines' known as ribosomes string together amino acids into long, linear chains. There are more than two hundreds types of amino acids, but only twenty of them exists in living organism. Amino acids links between each other by peptide bonds forming polypeptide chain, which sequence is is defined by a gene with genetic code. A typical protein contains 200-300 amino acids but some are much smaller (the smallest are often called peptides) and some much larger (the largest to date is titin a protein found in skeletal and cardiac muscle; it contains 26,926 amino acids in a single chain!). Number of different proteins, which it is possible to produce from 20 types of amino acids is enormous. For example for 10 amino acid sequence it is possible to have 2010 different sequences, which is approximately equal to 1013 or 10 trillions of different structures.

Formation of polypeptide chain is just a first step toward to completely functional protein. Like shoelaces, these chains loop about each other in a variety of ways (i.e., they fold) forming rigid globule. But, as with a shoelace, only one of these many ways allows the protein to function properly. Yet lack of function is not always the worst scenario. For just as a hopelessly knotted shoelace could be worse than one that won't stay tied, too much of a misfolded protein could be worse than too little of a normally folded one. This is because a misfolded protein can actually poison the cells around it. Recent discoveries show that Alzheimer's disease, cystic fibrosis, mad cow disease, even many cancers result from protein folding gone wrong.

Although it is possible to deduce the primary structure of a protein from a gene's sequence, its tertiary structure cannot be determined (although it should become possible to make predictions when more tertiary sequences are submitted to databases). It can only be determined by complex experimental analyses and, at present, this information is only known for about 10% of proteins. It is therefore not yet known how an amino-acid chain folds into its tertiary structure in the short time scale (fractions of a second) that occurs in the cell. So, there is a huge gap in our knowledge of how we move from protein sequence to function in living organisms: the line of sight from the genetic blueprint for a protein to its biological function is blocked by the impenetrable jungle of protein folding, and some researchers believe that clearing this jungle is the most important task in biochemistry and other field of science at present.

One of the most important results in understanding the process of protein folding was a thought-provoking experiment that was carried out by Christian Anfinsen and colleagues in the early 1960s. They investigated a protein called ribonuclease, which they isolated from the pancreatic tissue of cattle. Ribonuclease can be denatured by adding certain chemicals or by heat. In various studies, Anfinsen showed that this denaturation process could be completely reversed by removing these denaturing chemicals or by lowering the temperature. Anfinsen argued that the amino-acid sequence determines the shape of a protein. He received the Nobel Prize in Chemistry in 1972 for this finding.

If Anfinsen findings were true, then one more question becomes important. How do proteins find the right conformation out of the simply endless number of potential three-dimensional forms that it could randomly fold into? Cyrus Levinthal calculated in 1969 that finding the strongest attraction by simple trial and error would be impossible. He said that even if a protein only consisted of 100 amino acids and each of these flexible residues could only take on two different spatial orientations, the protein could theoretically adopt as many as 1030 possible conformations. Assuming a protein could try out 100 billion different conformations per second, it would still take 100 billion years to try all possibilities. So, Levinthal suggested that nature must have devised more effective methods to achieve this - and postulated the existence of defined sets of folding pathways by which protein folding can take place rapidly.

However, we now know that such fixed protein folding pathways do not seem to exist. Various protein folding pathways that have been investigated experimentally and theoretically in recent years have thrown up interesting hypotheses, but have remained hard to prove in working models. In the case of proteins of less than 100 amino acids, only two levels of folding can be observed, the unfolded protein (which occurs in numerous forms) and the finished, folded, functional protein. Beside two stages existing in short proteins in large one the third step or intermediate state exists. This state is known as molten globule. However, there is no evidence about whether these intermediates are formed en route to the correct folding pattern.

As with all processes in nature, protein folding also needs energy - the process has to obey the laws of thermodynamics. A protein always folds so that it achieves the lowest possible energy - just as we always try to adopt the most comfortable position, in which we need to move about least, when going to sleep. It is thought that this is achieved by using an energy gradient or 'funnel' along the path from the random tangle to the folded protein. Alan Fersht of Cambridge University used the following analogy to illustrate this model: if you blindfold a golfer and let him hit the ball in any direction he likes, the probability that he will hole the ball is almost infinitesimal. The same is true of a protein finding the right form by chance. However, if all parts of the golf course slope toward the hole, which is at the lowest point in the area, even a blindfolded golfer has a good chance of finding the hole. So, fixed reaction pathways are not necessary, as each protein seeks out its natural shape through a funnel of declining energy; it can take many folding routes and still reach its target of the completed tertiary structure.

We now understand better than ever how protein folding takes place. And this, in turn, has given us a better understanding of the origin and course of diseases that are associated with defective protein folding. But why the normal folding of every protein always runs towards a predetermined goal and what this goal looks like - that is, what the instructions in the primary structure are that determine the correct tertiary structure - is still a great mystery, even though the number of proposed models has dramatically increased. However, our improved understanding of the route that a protein must take from its synthesis to the correct folded form already enables us to contemplate better treatments or even cures for diseases in which proteins have departed from the correct folding route.


(Published: 10.12.2008.)