B. Alberts, A. Johnson, J. Lewis и др. - Molecular Biology of The Cell (6th edition) (1120996), страница 53
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Once a three-dimensional structure has been determined for onemember of a protein family, evolutionary tracing allows biologists to determinebinding sites for the members of that family, providing a useful start in deciphering protein function.Proteins Bind to Other Proteins Through Several Types ofInterfacesProteins can bind to other proteins in multiple ways.
In many cases, a portionof the surface of one protein contacts an extended loop of polypeptide chain (aMBoC6 m3.39/3.36“string”) on a second protein (Figure 3–41A). Such a surface–stringinteraction,for example, allows the SH2 domain to recognize a phosphorylated polypeptideloop on a second protein, as just described, and it also enables a protein kinase torecognize the proteins that it will phosphorylate (see below).A second type of protein–protein interface forms when two α helices, one fromeach protein, pair together to form a coiled-coil (Figure 3–41B).
This type of protein interface is found in several families of gene regulatory proteins, as discussedin Chapter 7.The most common way for proteins to interact, however, is by the precisematching of one rigid surface with that of another (Figure 3–41C). Such interactions can be very tight, since a large number of weak bonds can form between twosurfaces that match well. For the same reason, such surface–surface interactionscan be extremely specific, enabling a protein to select just one partner from themany thousands of different proteins found in a cell.stringsurfacesurface 1surface 2helix 2(A)SURFACE–STRINGhelix 1(B) HELIX–HELIX(C)SURFACE–SURFACEFRONTFigure 3–40 The evolutionary tracemethod applied to the SH2 domain.(A) Front and back views of a spacefilling model of the SH2 domain, withevolutionarily conserved amino acids on theprotein surface colored yellow, and thosemore toward the protein interior coloredred.
(B) The structure of one specific SH2domain with its bound polypeptide. Here,those amino acids located within 0.4 nmof the bound ligand are colored blue. Thetwo key amino acids of the ligand areyellow, and the others are purple. Note thehigh degree of correspondence between(A) and (B). (Adapted from O. Lichtarge,H.R. Bourne and F.E. Cohen, J. Mol. Biol.257:342–358, 1996. With permission fromElsevier; PDB codes: 1SPR, 1SPS.)Figure 3–41 Three ways in which twoproteins can bind to each other.
Onlythe interacting parts of the two proteinsare shown. (A) A rigid surface on oneprotein can bind to an extended loop ofpolypeptide chain (a “string”) on asecond protein. (B) Two α helices canbind together to form a coiled-coil.(C) Two complementary rigid surfacesoften link two proteins together. Bindinginteractions can also involve the pairing ofβ strands (see, for example, Figure 3–18).Chapter 3: Proteins138heavy chainVHVHhypervariable loopsNH2SSSSCH1 CH1SSSSSSVLSSSSSSSSSS SS SSVLCLCLCH2SSSSC H2CH3SSSSvariable domainof light chain (VL)disulfidebondCH3(A)constant domainof light chain (CL)COOH(B)Antibody Binding Sites Are Especially VersatileAll proteins must bind to particular ligands to carry out their various functions.The antibody family is notable for its capacity for tight, highly selective binding(discussed in detail in Chapter 24).Antibodies, or immunoglobulins, are proteins produced by the immune system in response to foreign molecules, such as those on the surface of an invading microorganism.
Each antibody binds tightly to a particular target molecule,thereby either inactivating the target molecule directly or marking it for destruction. An antibody recognizes its target (called an antigen) with remarkable specificity. Because there are potentially billions of different antigens that humansmight encounter, we have to be able to produce billions of different antibodies.Antibodies are Y-shaped molecules with two identical binding sites that arecomplementary to a small portion of the surface of the antigen molecule. Adetailed examination of the antigen-binding sites of antibodies reveals that theyMBoC6are formed from several loops of polypeptidechainm25.32/24.28that protrude from the endsof a pair of closely juxtaposed protein domains (Figure 3–42). Different antibodies generate an enormous diversity of antigen-binding sites by changing only thelength and amino acid sequence of these loops, without altering the basic proteinstructure.Loops of this kind are ideal for grasping other molecules.
They allow a largenumber of chemical groups to surround a ligand so that the protein can link to itwith many weak bonds. For this reason, loops often form the ligand-binding sitesin proteins.The Equilibrium Constant Measures Binding StrengthMolecules in the cell encounter each other very frequently because of their continual random thermal movements. Colliding molecules with poorly matchingsurfaces form few noncovalent bonds with one another, and the two moleculesdissociate as rapidly as they come together.
At the other extreme, when manynoncovalent bonds form between two colliding molecules, the association canpersist for a very long time (Figure 3–43). Strong interactions occur in cells whenever a biological function requires that molecules remain associated for a longtime—for example, when a group of RNA and protein molecules come together tomake a subcellular structure such as a ribosome.Figure 3–42 An antibody molecule.A typical antibody molecule is Y-shapedand has two identical binding sites forits antigen, one on each arm of the Y. Asexplained in Chapter 24, the protein iscomposed of four polypeptide chains (twoidentical heavy chains and two identicaland smaller light chains) held togetherby disulfide bonds.
Each chain is madeup of several different immunoglobulindomains, here shaded either blue or gray.The antigen-binding site is formed wherea heavy-chain variable domain (VH) anda light-chain variable domain (VL) comeclose together. These are the domains thatdiffer most in their sequence and structurein different antibodies.
At the end of eachof the two arms of the antibody molecule,these two domains form loops that bind tothe antigen (see Movie 24.5).PROTEIN FUNCTION139BBAthe surfaces of molecules A and B,and A and C, are a poor match andare capable of forming only a fewweak bonds; thermal motion rapidlybreaks them apartAACAACmolecule A randomly encountersother molecules (B, C, and D)DAADWe can measure the strength with which any two molecules bind to eachother. As an example, consider a population of identical antibody molecules thatsuddenly encounters a population of ligands diffusing in the fluid surroundingthem.
At frequent intervals, one of the ligand molecules will bump into the binding site of an antibody and form an antibody–ligand complex. The population ofantibody–ligand complexes will therefore increase,MBoC6but notwithout limit: overm3.42/3.39time, a second process, in which individual complexes break apart because ofthermally induced motion, will become increasingly important. Eventually, anypopulation of antibody molecules and ligands will reach a steady state, or equilibrium, in which the number of binding (association) events per second is preciselyequal to the number of “unbinding” (dissociation) events (see Figure 2–30).From the concentrations of the ligand, antibody, and antibody–ligand complexat equilibrium, we can calculate a convenient measure of the strength of binding—the equilibrium constant (K)—(Figure 3–44A).
This constant was describedin detail in Chapter 2, where its connection to free energy differences was derived(see p. 62). The equilibrium constant for a reaction in which two molecules (A andB) bind to each other to form a complex (AB) has units of liters/mole, and halfof the binding sites will be occupied by ligand when that ligand’s concentration(in moles/liter) reaches a value that is equal to 1/K.
This equilibrium constant islarger the greater the binding strength, and it is a direct measure of the free-energy difference between the bound and free states (Figure 3–44B). Even a change1A BdissociationA+Bdissociation rate = dissociation × concentrationrate constantof ABdissociation rate = koff [AB]2A+Bassociation rate =associationThe relationship betweenstandard free-energydifferences (ΔG°) andequilibrium constants (37°C)standardfree-energydifferenceof AB minus[AB]= K free energy[A][B]of A + B(liters/mole) (kJ/mole)equilibriumconstantA Bassociation × concentration × concentrationof Aof Brate constantassociation rate = kon [A] [B]31101021031041051061071081091010AT EQUILIBRIUM:association rate = dissociation ratekon [A] [B][AB][A][B](A)=konkoff=koff [AB]= K = equilibrium constant(B)0–5.9–11.9–17.8–23.7–29.7–35.6–41.5–47.4–53.4–59.4the surfaces of molecules A and Dmatch well and therefore can formenough weak bonds to withstandthermal jolting; they thereforestay bound to each otherFigure 3–43 How noncovalent bondsmediate interactions betweenmacromolecules (see Movie 2.1).Figure 3–44 Relating standardfree-energy difference (ΔG°) to theequilibrium constant (K).
(A) Theequilibrium between molecules A andB and the complex AB is maintained bya balance between the two opposingreactions shown in panels 1 and 2.Molecules A and B must collide if theyare to react, and the association rate istherefore proportional to the product of theirindividual concentrations [A] × [B]. (Squarebrackets indicate concentration.) As shownin panel 3, the ratio of the rate constantsfor the association and the dissociationreactions is equal to the equilibriumconstant (K) for the reaction (see also p.63). (B) The equilibrium constant in panel3 is that for the reaction A + B ↔ AB, andthe larger its value, the stronger the bindingbetween A and B.
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