Anti-GH1 monoclonal antibody, clone 9.G.34

The group A streptococcus Streptococcus pyogenes is the causative agent of a wide spectrum of invasive infections, including necrotizing fasciitis, scarlet fever and toxic shock syndrome. In the context of its carbohydrate chemistry, it is interesting that S. pyogenes (in this work strain M1 GAS SF370) displays a spectrum of oligosaccharide-processing enzymes that are located in close proximity on the genome but that the in vivo function of these proteins remains unknown. These proteins include different sugar transporters (SPy1593 and SPy1595), both GH125 alpha-1,6- and GH38 alpha-1,3-mannosidases (SPy1603 and SPy1604), a GH84 beta-hexosaminidase (SPy1600) and a putative GH2 beta-galactosidase (SPy1586), as well as SPy1599, a family GH1 'putative beta-glucosidase'. Here, the solution of the three-dimensional structure of SPy1599 in a number of crystal forms complicated by unusual crystallographic twinning is reported. The structure is a classical (beta/alpha)(g)-barrel, consistent with CAZy family GH1 and other members of the GH-A clan. SPy1599 has been annotated in sequence depositions as a beta-glucosidase (EC 3.2.1.21), but no such activity could be found; instead, three-dimensional structural overlaps with other enzymes of known function suggested that SPy1599 contains a phosphate-binding pocket in the active site and has possible 6-phospho-beta-glycosidase activity. Subsequent kinetic analysis indeed showed that SPy1599 has 6-phospho-beta-glucosidase (EC 3.2.1.86) activity. These data suggest that SPy1599 is involved in the intracellular degradation of 6-phosphoglycosides, which are likely to originate from import through one of the organism's many phosphoenolpyruvate phosphotransfer systems (PEP-PTSs).

Background: Ellagitannase (Ellagitannin acyl hydrolase) is an inducible enzyme with great potential use in food industry since allows the ellagic acid release from ellagitannins. Objective: In this work, ellagitannase was produced by the fungus Aspergillus niger GH1 in solid slate fermentation using polyurethane foam as solid support and pomegranate husk ellagitannins as sole carbon source and ellagitannase inducer and an initial approach to the enzymatic reaction conditions was reached. Materials and Methods: Ellagitannase was produced by Aspergillus niger GH1 in solid state fermentation and the ideal reaction conditions for ellagitannase activity based on ellagic acid quantification as ellagitannins biotransformation product by high performance liquid chromatographic are reported. Results:The enzyme ideal reaction conditions were: substrate concentration of 1 mg.mL(-1), 60 degrees C and pH 5.0, during 10 min of reaction. The kinetic enzyme constants (V-max = 30.34 niM.mL(-1).min(-1) and K-m = 1.48 x 10(3) mM) using punicalagin as substrate were detennined. Conclusion: The assay was completed in a short time and may find application in future studies of ellagic acid production.