Glycoside Hydrolases

• Author: Stephen Withers
• Responsible Editor: Spencer Williams

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Overview

Glycoside hydrolases are enzymes that catalyze the hydrolysis of the glycosidic linkage of glycosides, leading to the formation of a sugar hemiacetal or hemiketal and the corresponding free aglycon. Glycoside hydrolases are also referred to as glycosidases. Glycoside hydrolases can catalyze the hydrolysis of O-, N- and S-linked glycosides.

Classification

Glycoside hydrolases can be classified in many different ways. The following paragraphs list several different ways, the utility of which depends on the context in which the classification is made and used.

Endo/exo

exo- and endo- refers to the ability of a glycoside hydrolase to cleave a substrate at the end (most frequently, but not always the non-reducing end) or within the middle of a chain. For example, most cellulases are endo-acting, whereas LacZ β-galactosidase from E. coli is exo-acting.

Enzyme Commission (EC) number

EC numbers are codes representing the Enzyme Commission number. This is a numerical classification scheme for enzymes, based on the chemical reactions they catalyze. Every EC number is associated with a recommended name for the respective enzyme. EC numbers do not specify enzymes, but enzyme-catalyzed reactions. If different enzymes (for instance from different organisms) catalyze the same reaction, then they receive the same EC number. A necessary consequence of the EC classification scheme is that codes can be applied only to enzymes for which a function has been biochemically identified. Additionally, certain enzymes can catalyze reactions that fall in more than one class. These enzymes must bear more than one EC number.

Mechanistic classification

Two reaction mechanisms are most commonly found for the retaining and inverting enzymes, as first outlined by Koshland and as described below.{Koshland, 1953 #59} However several interesting variations on these mechanisms have been found, and one fundamentally different mechanism, catalyzed by an NADH cofactor, has been discovered in recent years, as discussed below.

Sequence classification

Sequence classification methods require knowledge of at least part of the amino acid sequence for an enzyme. Algorithmic methods are then used to compare sequences, and in the case of the glycoside hydrolases, this has allowed their classification into more than 100 families. Each family (GH family) contains proteins that are related by sequence, and by corollary, fold. Sequence-based classification schemes allow classification of proteins for which no biochemical evidence may have been obtained. An obvious shortcoming of sequence-based classifications are that they can only be applied to enzymes for which sequence information is available. As such sequence based classification methods are rather different (and in many ways complimentary) to the EC method discussed above.

Mechanism

Two reaction mechanisms are most commonly found for the retaining and inverting enzymes, as first outlined by Koshland and as described below [1]. However several interesting variations on these mechanisms have been found, and one fundamentally different mechanism, catalyzed by an NADH cofactor, has been discovered in recent years.

Inverting glycoside hydrolases

Hydrolysis of a glycoside with net inversion of anomeric configuration is generally achieved via a one step, single-displacement mechanism involving oxocarbenium ion-like transition states, as shown below. The reaction occurs with acid/base assistance from two amino acid side chains, normally glutamic or aspartic acids, that are typically located 6-11 A apart[2].

Retaining glycoside hydrolases

Hydrolysis with net retention of configuration is most commonly achieved via a two step, double-displacement mechanism involving a covalent glycosyl-enzyme intermediate, as is shown in Figure 3. Each step passes through an oxocarbenium ion-like transition state. Reaction occurs with acid/base assistance provided by two amino acid side chains, typically glutamate or aspartate, located 5.5 A apart. In the first step (often called the glycosylation step), one residue plays the role of a nucleophile, attacking the anomeric centre to displace the aglycon and form a glycosyl enzyme intermediate. At the same time the other residue functions as an acid catalyst and protonates the glycosidic oxygen as the bond cleaves. In the second step (known as the deglycosylation step), the glycosyl enzyme is hydrolyzed by water, with the other residue now acting as a base catalyst deprotonating the water molecule as it attacks. The pKa value of the acid/base group cycles between high and low values during catalysis to optimize it for its role at each step of catalysis [3].

References

1. Koshland, D. (1953) Biol. Rev. 28, 416.

2. McCarter JD and Withers SG. (1994). Mechanisms of enzymatic glycoside hydrolysis. Curr Opin Struct Biol. 1994;4(6):885-92. DOI:10.1016/0959-440x(94)90271-2 |
3. McIntosh LP, Hand G, Johnson PE, Joshi MD, Körner M, Plesniak LA, Ziser L, Wakarchuk WW, and Withers SG. (1996). The pKa of the general acid/base carboxyl group of a glycosidase cycles during catalysis: a 13C-NMR study of bacillus circulans xylanase. Biochemistry. 1996;35(31):9958-66. DOI:10.1021/bi9613234 |