4bce
crystal structure of Ttb-gly N282T mutantcrystal structure of Ttb-gly N282T mutant
Structural highlights
FunctionPublication Abstract from PubMedA large number of retaining glycosidases catalyze both hydrolysis and transglycosylation reactions, but little is known about what determines the balance between these two activities (transglycosylation/hydrolysis ratio). We previously obtained by directed evolution the mutants F401S and N282T of Thermus thermophilus beta-glycosidase (Ttbeta-gly, glycoside hydrolase family 1 (GH1)), which display a higher transglycosylation/hydrolysis ratio than the wild-type enzyme. In order to find the cause of these activity modifications, and thereby set up a generic method for easily obtaining transglycosidases from glycosidases, we determined their X-ray structure. No major structural changes could be observed which could help to rationalize the mutagenesis of glycosidases into transglycosidases. However, as these mutations are highly conserved in GH1 beta-glycosidases and are located around the -1 site, we pursued the isolation of new transglycosidases by targeting highly conserved amino acids located around the active site. Thus, by single-point mutagenesis on Ttbeta-gly, we created four new mutants that exhibit improved synthetic activity, producing disaccharides in yields of 68-90% against only 36% when native Ttbeta-gly was used. As all of the chosen positions were well conserved among GH1 enzymes, this approach is most probably a general route to convert GH1 glycosidases into transglycosidases. Semi-rational approach for converting a GH1 beta-glycosidase into a beta-transglycosidase.,Teze D, Hendrickx J, Czjzek M, Ropartz D, Sanejouand YH, Tran V, Tellier C, Dion M Protein Eng Des Sel. 2014 Jan;27(1):13-9. doi: 10.1093/protein/gzt057. Epub 2013 , Nov 27. PMID:24287187[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. See AlsoReferences
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