Structure and activity of the Streptococcus pyogenes family GH1 6-phospho β-glycosidase, Spy1599

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 -1,6- and GH38 -1,3-mannosidases (SPy1603 and SPy1604), a GH84 -hexosaminidase (SPy1600) and a putative GH2 -galactosidase (SPy1586), as well as SPy1599, a family GH1 `putative -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 (/)8-barrel, consistent with CAZy family GH1 and other members of the GH-A clan. SPy1599 has been annotated in sequence depositions as a -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--glycosidase activity. Subsequent kinetic analysis indeed showed that SPy1599 has 6-phospho--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)

Enhancement of poly(L-lactide)-degrading enzyme production by Laceyella sacchari LP175 using agricultural crops as substrates and its degradation of poly(L-lactide) polymer

Optimization of the medium for poly(L-lactide) (PLLA)-degrading enzyme production in shake flask experiments was investigated using agricultural products as substrates. Cassava chips and soybean meal were the best carbon and nitrogen sources, respectively, as determined by the one-factor-at-a-time method. Enzyme production was significantly enhanced by the addition of phosphates, but was suppressed by the addition of ammonium salt. The maximum enzyme production, 65.5 U/mL, was obtained from the optimized medium consisting of 4.64 g/L cassava chips, 1.53 soybean meal and 031 g/L PLLA powder using central composite design in the basal medium. The optimal physical factors in 3 L airlift fermenter were 50 degrees C, pH 7.0 and 0.5 vvm aeration rate for 18 h, yielded 94.4 U/mL. The crude enzyme was shown to be able to hydrolyze PLLA powder (91%) at 50 degrees C in 72 h which showed high efficiency for recycling of PLLA polymer and reducing the global environmental problem