Lodish H. - Molecular Cell Biology (5ed, Freeman, 2003) (794361), страница 32
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Antibodies act as specific sensors for antigens,forming antibody–antigen complexes that initiate a cascadeof protective reactions in cells of the immune system.All antibodies are Y-shaped molecules formed fromtwo identical heavy chains and two identical light chains(Figure 3-15a). Each arm of an antibody molecule containsa single light chain linked to a heavy chain by a disulfidebond. Near the end of each arm are six highly variable loops,called complementarity-determining regions (CDRs), whichform the antigen-binding sites. The sequences of the six loopsare highly variable among antibodies, making them specificfor different antigens.
The interaction between an antibodyand an epitope in an antigen is complementary in all cases;that is, the surface of the antibody’s antigen-binding sitephysically matches the corresponding epitope like a glove74CHAPTER 3 • Protein Structure and Function▲ FIGURE 3-15 Antibody structure and antibody-antigeninteraction. (a) Ribbon model of an antibody. Every antibodymolecule consists of two identical heavy chains (red) and twoidentical light chains (blue) covalently linked by disulfide bonds.(b) The hand-in-glove fit between an antibody and an epitope onits antigen—in this case, chicken egg-white lysozyme. Regions(Figure 3-15b). The intimate contact between these two surfaces, stabilized by numerous noncovalent bonds, is responsible for the exquisite binding specificity exhibited by anantibody.The specificity of antibodies is so precise that they candistinguish between the cells of individual members of aspecies and in some cases can distinguish between proteinsthat differ by only a single amino acid.
Because of their specificity and the ease with which they can be produced, antibodies are highly useful reagents in many of the experimentsdiscussed in subsequent chapters.Enzymes Are Highly Efficient and SpecificCatalystsIn contrast with antibodies, which bind and simply presenttheir ligands to other components of the immune system, enzymes promote the chemical alteration of their ligands,called substrates. Almost every chemical reaction in the cellis catalyzed by a specific enzyme.
Like all catalysts, enzymesdo not affect the extent of a reaction, which is determined bythe change in free energy G between reactants and products(Chapter 2). For reactions that are energetically favorable(G), enzymes increase the reaction rate by lowering theactivation energy (Figure 3-16). In the test tube, catalystssuch as charcoal and platinum facilitate reactions but usuallyonly at high temperatures or pressures, at extremes of highwhere the two molecules make contact are shown as surfaces.The antibody contacts the antigen with residues from all itscomplementarity-determining regions (CDRs). In this view, thecomplementarity of the antigen and antibody is especiallyapparent where “fingers” extending from the antigen surface areopposed to “clefts” in the antibody surface.or low pH, or in organic solvents.
As the cell’s protein catalysts, however, enzymes must function effectively in aqueousenvironment at 37C, 1 atmosphere pressure, and pH6.5–7.5.Two striking properties of enzymes enable them to function as catalysts under the mild conditions present in cells:their enormous catalytic power and their high degree ofspecificity. The immense catalytic power of enzymes causesthe rates of enzymatically catalyzed reactions to be 106–1012times that of the corresponding uncatalyzed reactions underotherwise similar conditions. The exquisite specificity ofenzymes—their ability to act selectively on one substrate or asmall number of chemically similar substrates —is exemplified by the enzymes that act on amino acids.
As noted inChapter 2, amino acids can exist as two stereoisomers, designated L and D, although only L isomers are normally foundin biological systems. Not surprisingly, enzyme-catalyzed reactions of L-amino acids take place much more rapidly thando those of D-amino acids, even though both stereoisomersof a given amino acid are the same size and possess the sameR groups (see Figure 2-12).Approximately 3700 different types of enzymes, each ofwhich catalyzes a single chemical reaction or set of closely related reactions, have been classified in the enzyme database.Certain enzymes are found in the majority of cells becausethey catalyze the synthesis of common cellular products (e.g.,proteins, nucleic acids, and phospholipids) or take part in the3.3 • Enzymes and the Chemical Work of CellsTransition state(uncatalyzed)Free energy, G∆GuncatTransition state(catalyzed)∆GcatReactantsProductsProgress of reaction▲ FIGURE 3-16 Effect of a catalyst on the activation energyof a chemical reaction.
This hypothetical reaction pathwaydepicts the changes in free energy G as a reaction proceeds. Areaction will take place spontaneously only if the total G of theproducts is less than that of the reactants (G). However, allchemical reactions proceed through one or more high-energytransition states, and the rate of a reaction is inverselyproportional to the activation energy (G‡), which is thedifference in free energy between the reactants and the highestpoint along the pathway. Enzymes and other catalysts acceleratethe rate of a reaction by reducing the free energy of thetransition state and thus G‡.production of energy by the conversion of glucose and oxygen into carbon dioxide and water. Other enzymes are present only in a particular type of cell because they catalyzechemical reactions unique to that cell type (e.g., the enzymesthat convert tyrosine into dopamine, a neurotransmitter, innerve cells).
Although most enzymes are located within cells,some are secreted and function in extracellular sites such asthe blood, the lumen of the digestive tract, or even outsidethe organism.The catalytic activity of some enzymes is critical to cellular processes other than the synthesis or degradation of molecules. For instance, many regulatory proteins and intracellularsignaling proteins catalyze the phosphorylation of proteins,and some transport proteins catalyze the hydrolysis of ATPcoupled to the movement of molecules across membranes.An Enzyme’s Active Site Binds Substratesand Carries Out CatalysisCertain amino acid side chains of an enzyme are importantin determining its specificity and catalytic power.
In the native conformation of an enzyme, these side chains arebrought into proximity, forming the active site. Active sitesthus consist of two functionally important regions: one thatrecognizes and binds the substrate (or substrates) and another that catalyzes the reaction after the substrate has been75bound.
In some enzymes, the catalytic region is part of thesubstrate-binding region; in others, the two regions are structurally as well as functionally distinct.To illustrate how the active site binds a specific substrateand then promotes a chemical change in the bound substrate,we examine the action of cyclic AMP–dependent protein kinase, now generally referred to as protein kinase A (PKA).This enzyme and other protein kinases, which add a phosphate group to serine, threonine, or tyrosine residues in proteins, are critical for regulating the activity of many cellularproteins, often in response to external signals. Because theeukaryotic protein kinases belong to a common superfamily, the structure of the active site and mechanism of phosphorylation are very similar in all of them.
Thus proteinkinase A can serve as a general model for this important classof enzymes.The active site of protein kinase A is located in the 240residue “kinase core” of the catalytic subunit. The kinasecore, which is largely conserved in all protein kinases, is responsible for the binding of substrates (ATP and a target peptide sequence) and the subsequent transfer of a phosphategroup from ATP to a serine, threonine, or tyrosine residuein the target sequence.
The kinase core consists of a large domain and small one, with an intervening deep cleft; the activesite comprises residues located in both domains.Substrate Binding by Protein Kinases The structure of theATP-binding site in the catalytic kinase core complements thestructure of the nucleotide substrate. The adenine ring of ATPsits snugly at the base of the cleft between the large and thesmall domains. A highly conserved sequence, Gly-X-Gly-XX-Gly-X-Val (X can be any amino acid), dubbed the “glycinelid,” closes over the adenine ring and holds it in position (Figure 3-17a). Other conserved residues in the binding pocketstabilize the highly charged phosphate groups.Although ATP is a common substrate for all protein kinases, the sequence of the target peptide varies among different kinases.
The peptide sequence recognized by proteinkinase A is Arg-Arg-X-Ser-Y, where X is any amino acid andY is a hydrophobic amino acid. The part of the polypeptidechain containing the target serine or threonine residue isbound to a shallow groove in the large domain of the kinasecore. The peptide specificity of protein kinase A is conferredby several glutamic acid residues in the large domain, whichform salt bridges with the two arginine residues in the target peptide. Different residues determine the specificity ofother protein kinases.The catalytic core of protein kinase A exists in an “open”and “closed” conformation (Figure 3-17b). In the open conformation, the large and small domains of the core region areseparated enough that substrate molecules can enter andbind.
When the active site is occupied by substrate, the domains move together into the closed position. This change intertiary structure, an example of induced fit, brings the target peptide sequence sufficiently close to accept a phosphate76CHAPTER 3 • Protein Structure and Function(a)Glycine lidSmall domainTargetpeptideNucleotidebindingpocketLarge domain(b)Glycine lidSmalldomainActive siteLargedomainthe active site. In the open position, ATP can enter and bindthe active site cleft; in the closed position, the glycine lid prevents ATP from leaving the cleft.
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