Enzyme: Number of Polypeptide and Number of Binding Sites (With Diagram)

Many enzymes, including lysozyme and ribonuclease, are composed of a single polypeptide chain.

Others consist of two or more chains. For example, the en­zyme glycogen phosphorylase consists of two polypep­tide chains; fumarase (a Krebs cycle enzyme) contains four polypeptide chains; and aspartate transcarbamy­lase (involved in the metabolic pathway leading to the synthesis of cytidine triphosphate) consists of 12 polypeptide chains.

Each of the enzyme’s constituent polypeptide chains is referred to as a subunit, the subunits being held together by electrostatic interac­tions, hydrogen bonds, van der Waals interactions, or other noncovalent forces that provide and stabilize protein quaternary structure.

Enzymes may have two separated, functionally different binding sites. One type of site, the active site, binds the substrate of the enzyme and possesses catalytic activity, whereas the other type of site, the allosteric (allo, “other” + steric, “space”) or regulatory site, lacks catalytic activity and binds an effector molecule. Such enzymes are usually referred to as allosteric enzymes.

Depending on the enzyme, active and allosteric sites may be on the same polypep­tide or on separate subunits. Effector molecules that inhibit enzyme activity (as in feedback inhibition) are called negative effectors.

In some cases, binding of an effector molecule enhances enzyme activity, and these molecules are known as positive effectors. A single enzyme may possess regulatory sites capable of bind­ing either negative or positive effectors; these effec­tors may compete with each other for the same regu­latory site or be bound at separate regulatory sites.

In some enzymes, two or more subunits may each possess a catalytic site, and substrate binding to the catalytic site on one subunit may influence substrate binding at another catalytic site. Such enzymes ex­hibit cooperativity—a phenomenon with the reversible oxygenation of he­moglobin. Enzymes that exhibit cooperativity and allosteric enzymes do not obey con­ventional Michaelis-Menten kinetics.

Model for Allosteric Enzyme Function:

A simple scheme depicting the influence of positive and negative effectors on allosteric enzyme activity is given in Figure 8-23, in which substrates and effec­tors and active and allosteric binding sites are repre­sented as geometric shapes. The active and regula­tory sites of the enzyme (in the absence of bound effectors) are depicted as circular areas.

The binding of a positive effector (hexagon) induces a conforma­tional change in the enzyme molecule at the regula­tory site (symbolized as a change from circular to hex­agonal shape). The conformational shape change is transmitted through the molecule (zigzag arrow) until it reaches and alters the active site in such a way that the substrate is bound more readily than in the absence of such activation.

In Figure 8-23, the altera­tion of the active site is represented as a change from circular to square shape. Binding of a negative effec­tor (diamond) induces a different type of conforma­tional change in the regulatory site (circular to dia­mond shape), which is transmitted through the enzyme’s structure such that the resulting change at the active site (circular to triangular shape) prevents subsequent substrate binding. The negative effector may, of course, be the end product of the pathway in­volving this enzyme.

The behavior of aspartate transcarbamylase, dis­cussed earlier in connection with feedback inhibition, serves as a good example of the mechanism depicted in Figure 8-23. Although aspartate transcarbamylase activity is inhibited by the end product CTP (i.e., CTP is a negative effector), ATCase activity is stimulated by ATP (i.e., ATP acts as a positive effector). It ap­pears that CTP and ATP compete for the same al­losteric sites on the enzyme.

The model of Figure 8-23 deliberately fails to stipu­late whether the allosteric enzyme is composed of one or more than one polypeptide chain, whether more than one active and regulatory site are present, or whether the sites are on the same or on different sub- units. Although all of these possibilities exist, most al­losteric enzymes consist of several polypeptide sub- units with active and regulatory sites on separate sub- units.

In the case of ATCase, there are 12 polypeptide chains—6 catalytic (C) chains (molecular weights of 34,000) and 6 regulatory (R) chains (molecular weights of 17,000). Each C chain contains one active site for aspartate and each R chain one regulatory site. Table 8-5 lists some of the allosteric enzymes, their substrates, and negative and positive effectors.