Enzyme Specificity and Selectivity

Abstract

Enzymes are exquisitely selective catalysts, capable of choosing a single substrate from a sea of similar compounds. Importantly, specificity is most manifest in the rate that a substrate reacts rather than the affinity of substrate binding. Specificity arises from the three‐dimensional structure of the enzyme‐active site, which is complementary to the transition state of the reaction. In some cases, a good substrate induces an active conformation that is not available to a poor substrate. Enzymes can also contain a second ‘proofreading’ site that further increases selectivity. Specificity is also evident in the way that enzymes control the decomposition of unstable intermediates, restricting their conformation so that the reaction is channelled down one pathway, to yield a single product. Lastly, it is important to recognize that enzyme selectivity is not absolute; the optimization of ‘promiscuous’ activities is an efficient route to the evolution of new enzymes.

Key concepts

  • Specificity is most manifest in the rate that a substrate reacts rather than the affinity of substrate binding.

  • The value of kcat/Km is used to assess specificity.

  • The enzyme‐active site is complementary to the transition state, with key residues perfectly aligned to promote the reaction.

  • A good substrate can induce an active enzyme conformation that is not accessed by a poor substrate.

  • Some enzymes contain additional active sites that catalyse proofreading reactions, further enhancing specificity.

  • Enzymes control reaction outcomes by restricting the conformation of substrates and high‐energy intermediates; this is called stereoelectronic control.

  • Enzyme specificity is not absolute; new enzymes evolve via the optimization of ‘promiscuous’ activities.

Keywords: lock and key; induced fit; stereoelectronic control; substrate synergism; promiscuous activity

Figure 1.

The hydrolysis of peptide bonds. (a) Acid‐catalysed peptide hydrolysis is indiscriminate, producing amino acids, whereas trypsin cleaves only after Lys and Arg residues (only Lys is shown). (b) The active site of trypsin accommodates an l‐peptide such that the bond to be cleaved is aligned with the catalytic residues. Note that the active site is complementary to the transition state, providing hydrogen bonds to stabilize the oxyanion. (c) In contrast, a d‐peptide is misaligned, and no reaction occurs. (d) The active sites of chymotrypsin and elastase are complementary to their substrates.

Figure 2.

Fidelity in the DNA polymerase reaction. Discrimination at the level of incorporation favours the correct nucleotide by a factor of 104. The correct nucleotide triphosphate induces the active conformation and is efficiently incorporated into the growing DNA strand. The incorrect nucleotide triphosphate induces a less active conformation and is inefficiently incorporated. DNA polymerase also has a proofreading mechanism that selectively removes the incorrect nucleotide, also by a factor of 104. Together, these two mechanisms account for the extraordinary fidelity of DNA replication.

Figure 3.

The lactate dehydrogenase reaction. Pyruvate and NADH are aligned such that the pro‐S hydrogen is transferred less than 2 times in 108 turnovers.

Figure 4.

Stereoelectronic control. (a) The triose‐phosphate isomerase reaction. When the enediol is formed in solution, elimination of the phosphate occurs to produce methylglyoxal. On the enzyme, the phosphate is held in the plane of the double bond to prevent the elimination reaction. (b) Pyridoxal phosphate chemistry. The carbanion is formed by removal of the group perpendicular to the plane of the ring. (c) The biosynthesis of sesquiterpenes. Over 200 different compounds are formed from farnesyl diphosphate; all of the reaction starts with the formation of the same carbonium ion. Each synthase constrains the allylic carbonium ion such that a single product (or a limited set of products) is formed.

Figure 5.

The evolution of new enzymes from promiscuous activities. (a) Modern enzymes are ‘specialists’, having evolved to optimally catalyse a given reaction. Nevertheless, these specialists often retain low‐level ability to catalyse other reactions. The enzyme can easily evolve to optimize such ‘promiscuous’ activities, usually via a ‘generalist’ that efficiently catalyses both reactions. (b) This process is illustrated by the in vitro evolution of aspartate aminotransferase (AATase) into tyrosine aminotransferase (TATase); the native enzyme efficiently catalyses the AATase reaction with very little TATase activity (blue numbers); the evolved enzyme efficiently catalyses both reactions (red numbers). Reprinted from Khersonsky et al. (2006). Copyright (2006), with permission from Elsevier.

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Further Reading

Christianson DW (2009) Unearthing the roots of the terpenome. Current Opinion in Chemical Biology 12: 141–150.

Fersht AR (1998) Structure and Mechanism in Protein Science, 1st edn. New York: WH Freeman.

Hedstrom L (1996) Trypsin: a case study in the structural determinants of enzyme specificity. Biological Chemistry 377: 465–470.

Jencks WP (1980) Binding energy, specificity, and enzymatic catalysis: the Circe effect. Advances in Enzymology and Related Areas of Molecular Biology 43: 219–410.

Johnson KA (2008) Role of induced fit in enzyme specificity: a molecular forward/reverse switch. Journal of Biological Chemistry 283: 26297–26301.

Khersonsky O, Roodveldt C and Tawfik DS (2006) Enzyme promiscuity: evolutionary and mechanistic aspects. Current Opinion in Chemical Biology 10: 498–508.

Knowles JR (1991) To build an enzyme …. Philosophical Transactions of the Royal Society London Series B 332: 115–121.

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How to Cite close
Hedstrom, Lizbeth(Feb 2010) Enzyme Specificity and Selectivity. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0000716.pub2]