Bio Factsheet. Proteins and Proteomics. Number 340

Transcription

1 Number 340 Proteins and Proteomics Every living thing on the planet is composed of cells, and cells in turn are made of many types of molecules, including the biological molecules carbohydrates, lipids, and proteins. This Factsheet will look at proteins and protein structure. Introduction Proteins are molecules, which means they are considerably smaller than cells. If we were to scale up a cell to be the size of a typical student s bedroom (about x100,000 magnification), amylase a protein you have probably heard of would be about the size of a grain of sugar! So, proteins may be tiny in comparison to the cells in which they operate, but the roles they play are diverse and of critical importance. Proteins and amino acids Like many important biological molecules, proteins are polymers, meaning they are composed of many smaller molecules (monomers) that have been chemically bonded together. The monomers are called amino acids, and there are 20 different types found in cells (see Table 1). Scientists often use a three- or one-letter code to abbreviate the names of these amino acids. Amino acid Three letter code One letter code alanine ala A arginine arg R asparagine asn N aspartic acid asp D cysteine cys C glutamic acid glu E glutamine gln Q glycine gly G histidine his H isoleucine ile I leucine leu L lysine lys K methionine met M phenylalanine phe F proline pro P serine ser S threonine thr T tryptophan trp W tyrosine tyr Y valine val V Table 1 Amino acids and their abbreviations Amino acids all share common features; they are called amino acids because they all possess an amino group (-NH 2 ) and a carboxylic acid group (-COOH). The difference between each type of amino acid lies in the chemical group they have attached to their central carbon atom. In the generalised structure of an amino acid this variable region is represented with an R (see Figure 1). Figure 1 Generalised structure of an amino acid This group can be as small as a single hydrogen atom, or larger than a benzene ring. Proteins vary enormously in their size, structure, and function. Some are very small (the hormone ADH is a protein made of only nine amino acids linked together), whereas others can be enormous (one molecule of titin a protein found in muscle tissue contains around 30,000 amino acids!) The differences are determined by both the number of amino acid monomers and which types of amino acids are present. This is because the R-groups determine exactly how the chain of amino acids will fold up. The gene that codes for any given protein is actually dictating the order and types of amino acids that will be linked together. This is why mutations in DNA can result in altered proteins; the amino acid sequence changes as a result of the change in the DNA sequence. Levels of protein structure Scientists like to classify things, and protein structure is no different. The main scheme used considers protein structure in a hierarchical series of four levels, referred to as primary, secondary, tertiary and quaternary structures. Each level involves different types of chemical bond, and different parts of the amino acid chain. Primary structure Primary structure describes which amino acids are found in a protein, and the exact order in which they are linked. So, for example, the entire primary structure of the hormone ADH can be written out (using the three-letter code) as: Cys-Tyr-Phe-Glu-Asn-Cys-Pro-Arg-Gly 1

2 (ii) Beta-pleated sheets (β-pleated sheets) The long chains that form a protein are not rigid, and as a newly-synthesised polypeptide starts to fold, sometimes lengths of the chain will double back and line up next to each other, held in place by hydrogen bonds between main-chain atoms. These structures are known as beta-pleated sheets (see Figure 4). Figure 2 Peptide bond formation When amino acids have been incorporated into a chain, scientists refer to them as amino acid residues. So, how are the amino acid residues held together to form long chains? The answer is by peptide bonds, a type of covalent bond formed during a condensation reaction between the amino group of one amino acid, and the carboxyl group of another. Being a condensation reaction, a molecule of water is generated during the process (see Figure 2). This reaction occurs in a ribosome, during the process of translation. Two amino acids joined together are called a dipeptide, whereas a longer chain of amino acids is called a polypeptide. No matter how long a polypeptide chain is, it will always have one end with an amino group and one end with a carboxyl group. Proteins are not simply snaking chains of amino acids linked by peptide bonds. These long chains fold up in a variety of ways. The next level of structure, secondary structure, refers to how the polypeptide chain starts to fold up. Secondary structure Secondary structure does not involve peptide bonds; instead, hydrogen bonds between main-chain atoms are used (R-groups are not involved in secondary structure bonding). There are two main types of secondary structure alpha helices and betapleated sheets. (i) Alpha helices (α-helices) Sometimes part of a chain of amino acids will form a spiral structure called an alpha-helix. This structure is held in place by hydrogen bonds between the hydrogen and oxygen atoms around peptide bonds that are four amino acids away from each other in the chain (see Figure 3). The amino acids involved in forming an alpha helix have their R-groups pointing to the outside of the helix. Figure 3 Alpha helix Tertiary structure As secondary structures form in a polypeptide, they in turn will fold up against each other and adopt a more stable three-dimensional shape, which is held in place by the formation of yet more bonds. In tertiary structure, there are a variety of different bond types that can occur, and they generally involve the R-groups of the amino acid residues instead of main-chain atoms. The three main types (see Figure 5) are as follows: (i) Hydrogen bonds The R-groups of many amino acids contain functional groups that are able to form hydrogen bonds. Note that hydrogen bonds between R-groups form tertiary structure, but hydrogen bonds between mainchain atoms form secondary structures. (ii) Ionic bonds Some amino acids have acidic or basic R-groups, and when two such groups are in close enough proximity it is possible for them to form an ionic bond. (iii) Disulphide bridges These are a type of covalent bond that can only form between the R-groups of two cysteine residues, through oxidation of their thiol groups (-SH): -SH HS- -S-S- + 2H Figure 5 Bonds in tertiary structure Figure 4 Beta-pleated sheet 2

3 For some proteins, tertiary structure is the last stage in their folding, and there is no higher level of structure. For example, the protein ribonuclease consists of a single polypeptide chain of 124 amino acids (the primary structure), which folds to form several alpha helices and beta pleated sheets (the secondary structures). The whole shape of the protein (the tertiary structure) is then held in place by the formation of four disulphide bridges. Figure 6 is a stereogram of ribonuclease in which the secondary structures are visible. If you can cross your eyes and make the two images overlap you will see a 3D image, with three prominent alpha helices close to you and several beta-pleated sheets behind them. Example #1: Haemoglobin, a globular protein In a typical human red blood cell, there are around ¼ billion molecules of haemoglobin, each of which can reversibly bind to four oxygen molecules. Haemoglobin provides a good example of a protein that has all levels of protein structure, from primary up to quaternary. (i) Primary structure Adult haemoglobin is composed of two different types of polypeptide chain. They are called α (alpha) and β (beta) chains, and two of each are present in a single haemoglobin molecule. The alpha chain consists of 141 amino acid residues, all held together by peptide bonds. The beta chain is slightly larger, and contains 146 amino acid residues. The primary structure of the alpha chain can be written out as follows, using the three-letter amino acid abbreviations seen in Table 1: Figure 6 Ribonuclease structure Quaternary structure In the case of proteins like ribonuclease, one polypeptide chain (i.e. a single molecule) is all that is needed to form a functional protein. However, many proteins are formed from more than one polypeptide chain. When this happens, each separate chain will fold up, and then the folded polypeptides will bond together to form the functional protein. This represents the final level of structure, quaternary structure, and the bonds involved in holding the polypeptides together are exactly the same as the ones found in tertiary structure. Prosthetic groups and cofactors Sometimes a folded protein requires the presence of a non-protein substance (e.g. a vitamin or a metal ion) in order to become functional. These substances are called cofactors. For example, the protein amylase the enzyme responsible for digesting starch only functions when chloride ions (Cl - ) are present. These chloride ions are not physically attached to the amylase, but they are required for it to function correctly. When a cofactor is tightly-bound to a protein, it is called a prosthetic group. Proteins with prosthetic groups are referred to as conjugated proteins. If you have studied cell membrane structure you will have heard of glycoproteins; these are proteins that are chemically attached to carbohydrate prosthetic groups, and they are the largest known group of conjugated proteins. val leu ser pro ala asp lys thr asn val lys ala ala try gly lys val gly ala his ala gly glu tyr gly ala glu ala leu glu arg met phe leu ser phe pro thr thr lys thr tyr phe pro his phe asp leu ser his gly ser ala gln val lys gly his gly lys lys val ala asp ala leu thr asn ala val ala his val asp asp met pro asn ala leu ser ala leu ser asp leu his ala his lys leu arg val asp pro val asp phe lys leu leu ser his cys leu leu val thr leu ala ala his leu pro ala glu phe thr pro ala val his ala ser leu asp lys phe leu ala ser val ser thr val leu thr ser lys tyr arg (ii) Secondary structure Each chain folds and forms eight alpha helices, a type of secondary structure described earlier on. The linking regions between them are formed by the remaining amino acid residues. The alpha helices are stabilised by many hydrogen bonds between mainchain atoms. (iii) Tertiary structure The alpha helices fold and form a compact three-dimensional shape, the tertiary structure, which is held in place by additional bonds, including hydrogen bonds between R-group atoms. Each subunit also has a hydrophobic pocket that contains a haem group (an Fe 2+ -containing prosthetic group) which is held in place by a covalent bond (Figure 7). Remember! In adult haemoglobin, the two types of haemoglobin subunit are called alpha and beta, but confusingly this has nothing to do with the secondary structures that are present; scientists just like using Greek letters! Haemoglobin subunit composition actually changes during your early life; in a developing embryo, the haemoglobin contains two zeta and two epsilon subunits, and in foetal haemoglobin there are two alpha and two gamma subunits! Classifying proteins Proteins are an incredibly diverse group of biological molecules, and there are many ways in which they can be classified. Proteins are often described as being either globular or fibrous depending on their structure and physical properties. Globular proteins have a roughly spherical shape and are soluble, with hydrophilic R-groups facing outwards on their surface, whereas fibrous proteins generally form long insoluble strands, with a hydrophobic exterior surface. 3 Figure 7 Haemoglobin subunit structure (haem group in black)

4 340. Proteins and Proteomics Figure 8 Three views of haemoglobin quaternary structure Proteomics Remember! The triple helix of a collagen molecule should not be confused with the alpha helices sometimes found in protein secondary structure. The triple helix is more extended and does not have the pattern of hydrogen bonding found in an alpha helix. Also, remember that an alpha helix is a single polypeptide chain that forms a tight coil, not three separate polypeptide chains twisted together! An interesting feature of collagen molecules is that many of them can bond together by forming crosslinks to make much larger structures called collagen fibrils. These fibrils can bundle together to make even larger collagen ibres (Figure 10). You probably know the term genome, which is used to describe all the DNA found in a set of chromosomes of a particular cell or organism. The term proteome might be less familiar, but the principle is the same; it means all of the proteins produced by a particular cell, tissue, organ or organism. Proteomics is a branch of biotechnology concerned with the study of proteomes. (iv) Quaternary structure A complete molecule of adult haemoglobin contains two α subunits and two β subunits. These are held together by hydrogen bonds and other non-covalent interactions to form the quaternary structure (Figure 8). Example #2: Collagen, a fibrous protein Collagen is a uniquely animal protein it is not found in any other kingdom of life. It is the most abundant protein in your body, and is used as a strong molecular glue to hold your cells, tissues and organs together. Tendons, ligaments, and connective tissue in general contain large quantities of collagen. Your proteome is determined by the way your genome is expressed. As proteins control cell activity, understanding the structure and function of proteins provides scientists with the information needed to tackle many important biological problems. Some examples of the applications of proteomics are: (i) Primary structure Designing drugs to target specific proteins involved in disease. Diagnosis of diseases by identifying proteins present in tissues. A single collagen molecule is formed from three long polypeptide chains. Each chain contains over 1000 amino acid residues. Determining which proteins are involved in biological processes. (ii) Secondary and tertiary structure The precise three-dimensional structures of proteins are determined using various methods, including x-ray crystallography, NMR, and more recently cryo-electron microscopy. The sequence data obtained by scientists is freely available on many online databases (e.g. the protein databank, and anyone in the world can search these databases and explore the structures of thousands of different proteins. The branch of science that deals with analysing the sequences of proteins (and genes) is called bioinformatics, and has been growing steadily over the past 40 years as computers have become more powerful and better software has been developed. Collagen molecules do not possess any organised secondary or tertiary structures, i.e. they do not have any α helices or β-pleated sheets, and the individual polypeptides do not fold up independently of each other. Instead, the three chains associate to form a quaternary structure. (iii) Quaternary structure The three polypeptide chains that form a single collagen molecule twist around each other to form an extended triple helix (see Figure 9). The chains pack closely together, and this is helped by the presence of large numbers of glycine residues in the chains. The amino acid glycine has a very small R group - a single hydrogen atom meaning there are fewer atoms to get in the way of the tightly grouped chains. Figure 9 The quaternary structure of collagen (only a short section of one molecule is shown) Figure 10 Collagen structure 4

5 Exam Questions 1. Carbonic anhydrase is an enzyme that is found in blood, liver and kidneys. The diagram below shows a molecular model of this enzyme. With reference to this diagram, and the parts labelled P and Q, explain the term secondary structure.. [3] (CIE) 2. Muscle cells contain globular and fibrous proteins. Compare and contrast the molecular structures of globular and fibrous proteins. [4] (Edexcel) 3. The primary amino acid sequence of a protein determines its final three-dimensional structure. The diagram shows two amino acids, cysteine and glycine. (i) The chemical groups used to form a peptide bond are A NH 2 and COOH B CH 2 and COOH C CH and CH 2 D NH 2 and CH [1] 5

6 (ii) The curliness of hair is the result of disulphide bonds between hair proteins. The more bonds, the curlier the hair. In recent years, hair straightening has become more popular. Thioglycolate is used by hairdressers before the hair is straightened. The process is completed using a second chemical to reverse the effect of the thioglycolate. Explain how this method keeps the hair straight. [2] (Edexcel) 4. The diagram below shows a molecule of haemoglobin. (a) State the inorganic ion present in the haem group. [1] (b) Using the diagram above, explain why this molecule is regarded as having a quaternary structure. [2] (WJEC) 5. (a) Amino acids form part of the structure of proteins. (i) State the name given to the sequence of amino acids in a protein molecule. [1] (ii) Draw the general structure of an amino acid molecule in the space below. [3] 6

7 (b) Collagen is an important fibrous protein which forms part of the wall of blood vessels. (i) State one property of collagen that makes it a useful component of blood vessel walls. [1] (ii) Describe the structure of the collagen molecule. [6] (OCR) Mark schemes 1. P is β-pleated sheet, Q is α-helix Accept if P and Q are identified by a description (1) Determined by, coiling/folding/sequence, of amino acids/polypeptide; A primary structure for sequence of amino acids (1) Stabilised/held/AW by hydrogen bonds (1) Between C = O and H N (of peptide bonds); A carbonyl/carboxyl group, and, amine/amino group (1) Ref to, parallel/anti-parallel, nature of β-pleated sheet. (1) (3 marks) 2. An answer which makes reference to four of the following: Both are chains of amino acids joined by peptide bonds (1) Both contain named bonds (holding molecule in its three-dimensional shape) (1) i.e. hydrogen bonds, disulphide bridges, ionic bonds. Globular proteins have hydrophilic groups on the outside, whereas fibrous proteins have hydrophobic groups on the outside (1) Allow converse. Globular have tertiary or quaternary structures whereas fibrous have little or no tertiary structure (1) Globular are folded into compact shapes, whereas fibrous have long chains (1) Allow globular being spherical and fibrous being long strands. (4 marks) 7

8 340 Proteins and Proteomics 3. (i) A (NH 2 and COOH) (1) (ii) An explanation that makes reference to the following: (a) Thioglycolate breaks disulphide bonds (1) (1 mark) The second chemical reforms the disulphide bonds, stopping the hair from becoming curly again (1) (a) Iron / Fe2 + ; (1 mark) (2 marks) (1 mark) (b) {Four polypeptide chains / two alpha and two beta subunits}; in tertiary form are {combined/joined} (2 marks) (i) Primary Structure (1 mark) Accept 1 structure Ignore polypeptide (ii) NH2 at one end (1 mark) Accept displayed structure of NH 2 / HNH COOH at opposite end (1 mark) Accept displayed structure of COOH if correct double bond shown (2 marks) (1 mark) C in centre (of a single amino acid) bonded (separately) to one R and one H (1 mark) Award only if the candidate has drawn a single amino acid molecule If R group not shown as R then award max 2 marks (as general structure asked for Q) Ignore labels (3 marks) (b) (i) strength / toughness / insolubility (1 mark) Mark the first answer. If the answer is correct and an additional answer is given that is incorrect or contradicts the correct answer then 0 marks. Accept strong/tough. Ignore flexible / inelastic. Ignore withstand pressure. (1 mark) (ii) One molecule of collagen is 3 polypeptide chains twisted around each other. Credit annotated diagrams unless contradicted by text. 1 peptide bonds, between amino acids / in polypeptide (1) 2 every 3rd amino acids, is same / glycine (1) Accept high proportion of / 35%, glycine / same amino acid 3 coil / twist / spiral / helix (1) Credit in context of single polypeptide or 3 polypeptides but do not credit a-helix in the context of a single polypeptide Ignore wound 4 left-handed (helix) (1) a-helix, which is left handed award mp4 but do not credit mp3 5 glycine / small R group, allows closeness / twisting (of polypeptide chains) (1) 6 three polypeptide chains (1) 7 hydrogen / H, bonds between (polypeptide) chains (1) Must be in correct context Do not credit H + / H 2 bonds 8 no / few hydrophilic (R) groups on outside (of molecule) (1) 9 (adjacent molecules joined by) crosslinks (1) Accept covalent bonds between adjacent molecules Do not credit in context of bonding between 3 polypeptides Ignore disulphide 10 crosslinks / ends of molecules, being staggered (1) 11 fibril (1) Ignore micro (6 marks) Acknowledgements: This Biology Factsheet was researched and written by Aaron Bridges and published in September 2017 by Curriculum Press. Biology Factsheets may be copied free of charge by teaching staff or students, provided that their school is a registered subscriber. No part of these Factsheets may be reproduced, stored in a retrieval system, or transmitted, in any other form or by any other means, without the prior permission of the publisher. ISSN