Riu In-Hyun, Shin Il-Soo, Do Su-Il
Department of Life Science, Laboratory of Functional Glycomics, Ajou University, San 5, Wonchon-dong, Youngtong-gu, Suwon City 443-749, Republic of Korea.
Biochem Biophys Res Commun. 2008 Jul 18;372(1):203-9. doi: 10.1016/j.bbrc.2008.05.034. Epub 2008 May 16.
cDNAs encoding three isoforms of OGT (ncOGT, mOGT, and sOGT) were expressed in Escherichia coli in which the coexpression system of OGT with target substrates was established in vivo. No endogenous bacterial proteins were significantly O-GlcNAcylated by any type of OGT isoform while co-expressed p62 and Sp1 were strongly O-GlcNAcylated by ncOGT. These results suggest that most of bacterial proteins appear not to be recognized as right substrates by mammalian OGT whereas cytosolic environments may supply UDP-GlcNAc enough to proceed to O-GlcNAcylation in E. coli. Under these conditions, sOGT was auto-O-GlcNAcylated whereas ncOGT and mOGT were not. Importantly, we found that when Sp1 was coexpressed, ncOGT can O-GlcNAcylate not only Sp1 but also many bacterial proteins. Our findings suggest that Sp1 may modulate the capability of target recognition of ncOGT by which ncOGT can be led to newly recognize bacterial proteins as target substrates, finally generating the O-glyco-bacteria. Our results demonstrate that the O-glyco-bacteria showed enhanced thermal resistance to allow cell survival at a temperature as high as 52 degrees C.
编码OGT三种同工型（ncOGT、mOGT和sOGT）的cDNA在大肠杆菌中表达，在大肠杆菌中体内建立了OGT与靶底物的共表达系统。在共表达时，没有任何一种OGT同工型能使内源性细菌蛋白发生显著的O-连接N-乙酰葡糖胺化，而共表达的p62和Sp1被ncOGT强烈O-连接N-乙酰葡糖胺化。这些结果表明，大多数细菌蛋白似乎不能被哺乳动物OGT识别为合适的底物，而胞质环境可能提供足够的UDP-GlcNAc以在大肠杆菌中进行O-连接N-乙酰葡糖胺化。在这些条件下，sOGT发生了自身O-连接N-乙酰葡糖胺化，而ncOGT和mOGT则没有。重要的是，我们发现当Sp1共表达时，ncOGT不仅能使Sp1发生O-连接N-乙酰葡糖胺化，还能使许多细菌蛋白发生O-连接N-乙酰葡糖胺化。我们的发现表明，Sp1可能调节ncOGT的靶标识别能力，通过这种能力，ncOGT可被引导重新将细菌蛋白识别为靶底物，最终产生O-糖基化细菌。我们的结果表明，O-糖基化细菌表现出增强的耐热性，使其能够在高达52摄氏度的温度下存活。
