Biochemistry 2.4 PDF

Protein ModificationsIntroductionAll proteins consist of at least one chain of amino acids linked together through. The sequence of amino acids in each chain determines the biological function carried out by that chain when the protein folds correctly. In addition to amino acid sequence, a protein\'...

Protein ModificationsIntroductionAll proteins consist of at least one chain of amino acids linked together through. The sequence of amino acids in each chain determines the biological function carried out by that chain when the protein folds correctly. In addition to amino acid sequence, a protein\'s ability to function often depends on chemical groups other than amino acids. This lesson discusses changes to proteins, both at the amino acid and the non--amino acid levels, that can affect protein function.2.4.01 MutationsGenes occasionally undergo random that can change the amino acid sequence of the corresponding protein. Consequently, the amino acid sequence for a given protein can vary from one species to another or between individuals in a species. Within a species, the most common amino acid sequence (ie, the sequence normally found in the wild) is called the wild-type form of that protein. Individuals with a sequence other than the wild-type sequence have a mutant form of the protein.Protein substitution mutations are typically indicated by the one-letter code of the amino acid from the wild-type protein, followed by its position in the amino acid sequence (in which the N-terminal residue is position 1), and the one-letter code of the new (mutant) amino acid. For example, a Q37L mutation means that in the wild-type protein the 37th amino acid is glutamine (Q), but in the mutant protein it has changed to leucine (L). Chapter 2: Peptides and Proteins80Concept Check 2.12The following image shows a mutation in a certain peptide sequence. What is the correct designation for this mutation?Different mutations have different effects on a protein\'s function. A mutation from one amino acid to another with similar chemical properties tends to have a smaller effect than mutations that significantly alter chemical properties. For example, the mutation K45R changes the amino acid at position 45 from one positively charged group (K, lysine) to another (R, arginine).This mutation is less likely to affect protein function than a K45E mutation, which changes a positive charge (K, lysine) to a negative charge (E, glutamate). Mutations that preserve chemical properties are called. Those that do not are called nonconservative.In addition to the type of mutation (eg, conservative vs. nonconservative), the location of a mutation within a protein impacts the effect of that mutation. For example, in flexible, unstructured regions, even nonconservative mutations are less likely to have a significant effect on protein function. In contrast, highly structured regions are more susceptible to disruption by mutation. Furthermore, regions that are critical to protein function may even be affected by conservative mutations. Examples of protein regions with different susceptibilities to mutation are shown in Figure 2.38. Chapter 2: Peptides and Proteins81Figure 2.38 Examples of the relation between location and the probable impact of mutations.Comparing the wild-type sequences for the same protein in different species reveals which amino acids are critical for function and which are not. When sequences of related proteins are properly aligned, the amino acids that are critical to structure and/or function are present in all species. These amino acids are said to be highly conserved.In many cases, the chemical properties are important, but any amino acid with those properties can serve (eg, serine and threonine are both acceptable). Amino acids at these positions are also said to be conserved but to a lesser extent than amino acids that must remain unaltered. Amino acids that vary significantly from one species to another are not conserved. Chapter 2: Peptides and Proteins82Concept Check 2.13The wild-type sequences of the protein hexokinase isolated from five different species were aligned. The following image shows a small portion of each sequence.If the amino acid residue at the left of each sequence is position 111, which position is the most critical for hexokinase structure and/or function? (Note: Sequence numbering is based on the sequence of species 1.)2.4.02 Post-translational ModificationsThe process of a ribosome synthesizing a protein is called. After a protein is translated, it may be altered by various enzymes acting on it. These enzymes often add chemical groups that are not amino acids to the protein. The resulting alterations are called post-translational modifications. Many types of post-translational modifications exist, including the following:PhosphorylationThis modification involves the transfer of a phosphate group, usually from ATP, to the side chain of an amino acid residue. Phosphorylation is facilitated by a class of enzymes called kinases (see Concept 4.2.03) and typically occurs on , , or residues. Each of these amino acids has a hydroxyl group that can act as a nucleophile to attack the group on ATP, yielding ADP and phosphoserine, phosphothreonine, or phosphotyrosine, respectively (Figure 2.39). Other amino acid side chains can also undergo phosphorylation, but this is much less common.Figure 2.39 General schematic of protein phosphorylation by ATP, facilitated by a kinase enzyme. Chapter 2: Peptides and Proteins83Phosphorylation often regulates protein function. The specific effect of phosphorylation depends on the protein being modified; some proteins are activated by phosphorylation, whereas others are inactivated.Concept Check 2.14The following graph shows the activity levels of two proteins under different conditions. What is the effect of phosphorylation on each protein?Phosphate groups are negatively charged and therefore typically exert their effect by adding negative charges to a protein. This added negative charge decreases the protein\'s and may induce conformational changes that alter the protein\'s ability to interact with its environment and with other molecules.Interestingly, mutation of a serine or threonine residue to glutamate or aspartate often produces the same effect as phosphorylation. For instance, if a protein is activated by phosphorylation of a specific serine residue, mutating that residue to aspartate can cause the protein to become constitutively active (ie, it cannot be inactivated). The negative charge of the aspartate residue has the same effect as the negative charge of a phosphate group, but aspartate cannot be removed by dephosphorylation. This type of mutation is known as a because it mimics the effect of a phosphate group. Chapter 2: Peptides and Proteins84GlycosylationMany proteins, particularly those synthesized on the , are modified by glycosylation (ie, the addition of carbohydrate groups). Glycosylation, shown in Figure 2.40, is catalyzed by glycosyltransferase enzymes. The added carbohydrate may consist of a single sugar or many sugars linked together. Additional information on carbohydrates can be found in Chapter 7.Glycosylation most commonly occurs in the lumen of the endoplasmic reticulum and the. The two most common types of glycosylation are N-linked and O-linked glycosylation, which are distinguished by the amino acid residues on which they take place.N-linked glycosylation occurs on the --NH2 groups of side chains. As the glycosylated protein moves from the ER to the Golgi, the complex carbohydrate may be modified by addition or removal of individual sugars. This process helps to direct proteins to the correct destinations (eg, lysosome, extracellular space), and may also facilitate protein folding and activity. O-linked glycosylation plays similar roles to N-linked glycosylation, but it occurs on the --OH groups of serine or threonine residues.Figure 2.40 Carbohydrates are added to serine, threonine, or asparagine residues in the ER.UbiquitinationThis modification involves two proteins: the target protein and ubiquitin. Therefore, unlike other post-translational modifications, ubiquitination involves addition of other amino acids (ie, those found in ubiquitin) to an existing protein. Chapter 2: Peptides and Proteins85Ubiquitination can occur when the N-terminus or a side chain (often ) of the target protein nucleophilically attacks the C-terminus of ubiquitin. Bonds between lysine side chains and ubiquitin C-termini are the most common linkages. This linkage forms an amide bond similar to the between amino acids in a protein backbone. However, because this bond forms between a side chain and a backbone group (rather than two backbone groups), it is called an rather than a peptide bond.Once one ubiquitin is added to a target protein, another ubiquitin can be added to the first. This process can be repeated multiple times to form a polyubiquitin chain. Ubiquitination serves many biological purposes, but the best characterized is ubiquitin\'s role in protein degradation. When cytosolic proteins are misfolded or no longer needed, they are commonly modified with a polyubiquitin tag, which then targets them to a protein complex called the proteasome (see Figure 2.41). This complex digests the protein into small peptide and amino acid fragments, which are then recycled to make other proteins.Figure 2.41 Polyubiquitin tags send proteins to the proteasome for degradation.AcetylationIn addition to modification by ubiquitin, lysine residues in a protein may also be modified by addition of an acetyl group. In these reactions, the side chain nitrogen of lysine acts as a nucleophile to attack the carbonyl carbon of the acetyl group in acetyl coenzyme A, producing acetylated lysine and free coenzyme A. In some cases, the N-terminus of a protein can act as the nucleophile in place of lysine.Many proteins undergo acetylation. Perhaps the best-known example is histone proteins (Figure 2.42), which bind DNA and are crucial to the structure of chromosomes. When unmodified, the positively charged lysine residues in a histone interact with the negatively charged DNA backbone. However, acetylation neutralizes the positive charge of lysine, releasing the DNA. This process is important in regulating which genes in a cell are transcribed to produce RNA and which are not. Chapter 2: Peptides and Proteins86Figure 2.42 Interactions between histones and DNA are disrupted by lysine acetylation.Lipidation is similar to acetylation in that a nucleophilic amino acid group attacks a carbonyl linked to a hydrocarbon. However, the acetyl group in acetylation contains a single carbon in its hydrocarbon chain, whereas lipidation involves much longer, more complex hydrocarbons. Lipidation occurs on lysine, cysteine, and serine side chains and is involved in many biological processes, including those carried out by certain important metabolic enzymes (see Chapter 12).ProteolysisUnlike other post-translational modifications, occurs by removing groups from proteins. This occurs when a protease enzyme hydrolyzes a peptide bond. Certain proteins must remain inactive until they are transported to the appropriate compartment of a cell or organism or until certain environmental conditions are met. To accomplish this, these proteins are synthesized in an inactive form called a proprotein. Upon reaching the correct environment, the proprotein undergoes proteolysis (also called cleavage) to convert it into its active form. Chapter 2: Peptides and Proteins872.4.03 Cofactors To function, many proteins require chemical groups other than amino acids. Any non--amino acid group required for protein function is called a cofactor. Cofactors may range in complexity from single metal ions (eg, Ca2+, Mg2+) to complex organic molecules. Organic cofactors are given the additional designation of coenzymes and often assist enzymes in carrying out their reactions. Coenzymes are commonly derived from.Some cofactors interact only transiently with their proteins. Other cofactors bind tightly or even covalently to their target proteins. Covalently bound cofactors can be thought of as post-translational modifications. These tightly bound cofactors effectively become part of the enzyme, and, as such, they are called.An important coenzyme found in many proteins, often as a prosthetic group, is heme. Heme is derived from the organic molecule porphyrin and contains an iron (Fe) atom at its center, bound to the four groups of heme. Heme is involved in several crucial oxidation-reduction reactions and in the transport and intracellular storage of oxygen. It has the characteristic structure shown in Figure 2.43.Figure 2.43 Structure of heme, with the porphyrin ring highlighted.All proteins are initially synthesized using only amino acids and must interact with any cofactors, including prosthetic groups, after translation (and often after folding). Some proteins that require prosthetic groups can be found without these groups in certain circumstances. In this form, these proteins are called apoproteins. Upon addition of the correct prosthetic group, an apoprotein is converted into a holoprotein. Only the holoprotein form can carry out the correct biological function.