Cloning, expression and characterization of a new endo-β-N-acetylglucosaminidase from Streptomyces alfalfae / 生物工程学报

Endo-β-N-acetylglucosaminidase is used widely in the glycobiology studies and industries. In this study, a new endo-β-N-acetylglucosaminidase, designated as Endo SA, was cloned from Streptomyces alfalfae ACCC 40021 and expressed in Escherichia coli BL21 (DE3). The purified recombinant Endo SA exhibited the maximum activity at 35 ºC and pH 6.0, good thermo/pH stability and high specific activity (1.0×10⁶ U/mg). It displayed deglycosylation activity towards different protein substrates. These good properties make EndoSA a potential tool enzyme and industrial biocatalyst.

Relationship of protein O-GlcNAcylation levels with antitumor effect of Taxol in breast cancer / 中国生化药物杂志

Objective To study effect of Taxol on protein O-GlcNAcylation levels and investigate whether protein O-GlcNAcylation levels can affect the sensitivity of breast cancer cells to Taxol.Methods Western blot analysis was performed to examine protein O-GlcNAcylation levels and the expression of enzymes related to O-GlcNAcylation biosynthesis in Taxol treated breast cancer cells.RT-qPCR was used to analyze the effects of Taxol on OGA and OGT mRNA expression in cancer cells.The sulforhodamine B colorimetric assay was used to determine the effect of alteration of protein O-GlcNAcylation on the anti-proliferation of Taxol in breast cancer cells by adding OGA inhibitor and OGT inhibitor, respectively.Results Taxol treatment enhanced protein O-GlcNAcylation levels in dose-and time-dependent manners in breast cancer cell MDA-MB-231 ( P <0.05 ) .Taxol increased the mRNA levels of OGT and OGA after MDA-MB-231 cells were treated for 24 h(P<0.05).As OGA inhibitors increased protein O-GlcNAcylation levels, the sensitivity of MDA-MB-231 to Taxol was increased.As OGT inhibitor decreased protein O-GlcNAcylation levels, the sensitivity of MDA-MB-231 to Taxol was reduced.Conclusion Taxol treatment can enhance protein O-GlcNAcylation levels and the changes of O-GlcNAcylation levels alter the sensitivity of breast cancer cell MDA-MB-231 to Taxol.