Physicochemical consequences of the perdeuteriation of glutathione S-transferase from S. japonicum

Glutathione S-transferase (GST) from Schistosoma japonicum has been prepare d in both normal protiated (pGST) and fully deuteriated (dGST) form by reco mbinant DNA technology, Electrospray mass spectrometry showed that the leve l of deuteriation in dGST was 96% and was homogeneous across the sample. Th is result is attributed to the use of a deuterium-tolerant host Escherichia coli strain in the preparation of the protein. 10 heteroatom-bound deuteri ums tin addition to the carbon-bound deuteriums) were resistant to exchange when dGST was incubated in protiated buffer. The physicochemical and biolo gical properties of the two proteins were compared. dGST was relatively les s stable to heat denaturation and to proteolytic cleavage than was pGST. Th e midpoint transition temperature fur pGST was 54.9 degreesC, whereas that for dGST was 51.0 degreesC. Static light-scattering measurements revealed t hat the association behavior of dGST is also different from that of pGST. T he perdeuteriated enzyme shows a tendency to associate into dimers of the f undamental dimer. This is in contrast with results that have been obtained for other perdeuteriated proteins in which perdeuteriation has been shown t o promote dissociation of aggregates. dGST showed a similar K, to pGST; sim ilar results had been obtained previously with bacterial alkaline phosphata se. However, whereas the alkaline phosphatase showed a reduced rate of cata lysis on deuteriation, dGST exhibited a slightly higher rate of catalysis t han pGST. It is clear that the bulk substitution of deuterium for protium h as significant effects on the properties of proteins. Until many more examp les have been studied, it will be difficult to predict these effects for an y given protein. Nevertheless, deuteriation represents an intriguing method of preparing functional analogs of recombinant proteins.