γ-Glutamyltranspeptidases: sequence, structure, biochemical properties, and biotechnological applications

Abstract

γ-Glutamyltranspeptidases (γ-GTs) are ubiquitous enzymes that catalyze the hydrolysis of γ-glutamyl bonds in glutathione and glutamine and the transfer of the released γ-glutamyl group to amino acids or short peptides. These enzymes are involved in glutathione metabolism and play critical roles in antioxidant defense, detoxification, and inflammation processes. Moreover, γ-GTs have been recently found to be involved in many physiological disorders, such as Parkinson's disease and diabetes. In this review, the main biochemical and structural properties of γ-GTs isolated from different sources, as well as their conformational stability and mechanism of catalysis, are described and examined with the aim of contributing to the discussion on their structure-function relationships. Possible applications of γ-glutamyltranspeptidases in different fields of biotechnology and medicine are also discussed.

Fig. 1

Multiple alignment of amino acid sequences of the small subunits of some γ-GTs. The amino acid sequences of γ-GT from H. sapiens (Swiss-Prot P19440), R. norvegicus (Swiss-Prot P07314), D. rerio (Swiss-Prot Q7T2A1), D. melanogaster (Swiss-Prot Q9VWT3), S. cerevisiae (Swiss-Prot Q05902) E. coli (Swiss-Prot P18956), H. pylori (Swiss-Prot O25743), B. subtilis (Swiss-Prot P54422), B. licheniformis (Swiss-Prot Q62WE3), G. thermodenitrificans (YP001127364.1), D. radiodurans (PIR: D75385), T. thermophylum (NCBI Reference Sequence: YP_144021.1) and T. acidophilum (NCBI Reference Sequence: NP_394454.1) are included. The alignment was performed by using the ClustalW method (http://www.ebi.ac.uk/Tools/msa/clustalw2/). The conserved threonine residues responsible for autoprocessing of the enzyme and the two strictly conserved glycine residues involved in binding of the γ-glutamyl moiety (T391, G483, and G484 in EcGT) are highlighted in yellow. The other residues responsible for substrate binding and catalytic activity of γ-GTs are highlighted in cyan (T409, N411, D433, S462, and S463 in EcGT). The lid loop extending towards the active site (spanning from P438 to G449 of EcGT) and absent in some γ-GTs is underlined. The secondary structure elements are shown above the alignment. The numbering scheme reported in the figure refers to the small subunit

Fig. 2

Diagram showing a schematic representation of the precursor (a) and mature (b) forms of EcGT. In panel b, the large subunit is shown in violet and the small subunit in yellow

Fig. 3

Proposed mechanism for autocatalytic processing of γ-GTs

Fig. 4

Superimposed structure of precursor (green) and mature forms of EcGT. In the mature enzyme, the large subunit is shown in violet and the small subunit in yellow

Fig. 5

Proposed reaction mechanism of γ-GTs

Fig. 6

Glutamate binding in the active site of EcGT. The stick model of l-glutamate, nucleophile Thr391 and of Tyr444 are shown. The large subunit is shown in violet and the small subunit in yellow

Fig. 7

Binding mode of l-glutamate in the catalytic pocket of EcGT. Carbon, nitrogen and oxygen atoms are colored cyan, blue and red. The stick models of residues involved in the ligand binding and enzyme reaction are shown