n-terminal deletion of peptiden-glycanase results in enhanced deglycosylation activity氨基端删除peptiden-glycanase增强deglycosylation活动的结果.pdfVIP

N-Terminal Deletion of Peptide:N-Glycanase Results in Enhanced Deglycosylation Activity 1 1 1 1 1 1,2 Shengjun Wang , Fengxue Xin , Xiaoyue Liu , Yuxiao Wang , Zhenyi An , Qingsheng Qi *, Peng George Wang1,2 1 State Key Laboratory of Microbial Technology, Shandong University, Jinan, China, 2 National Glycoengineering Research Center, Shandong University, Jinan, China Abstract Peptide:N-glycanase catalyzes the detachment of N-linked glycan chains from glycopeptides or glycoproteins by hydrolyzing the b-aspartylglucosaminyl bond. Peptide:N-glycanase in yeast binds to Rad23p through its N-terminus. In this study, the complex formed between Peptide:N-glycanase and Rad23p was found to exhibit enhanced deglycosylation activity, which suggests an important role for this enzyme in the misfolded glycoprotein degradation pathway in vivo. To investigate the role of this enzyme in this pathway, we made stepwise deletions of the N-terminal helices of peptide:N- glycanase. Enzymatic analysis of the deletion mutants showed that deletion of the N-terminal H1 helix (Png1p-DH1) enhanced the deglycosylation activity of N-glycanase towards denatured glycoproteins. In addition, this mutant exhibited high deglycosylation activity towards native glycoproteins. Dynamic simulations of the wild type and N-terminal H1 deletion mutant implied that Png1p-DH1 is more flexible than wild type Png1p. The efficient deglycosylation of Png1p-DH1 towards native and non-native glycoproteins offers a potential biotechnological application. Citation: Wang S, Xin F, Liu X, Wang Y, An Z, et al. (2009) N-Terminal Deletion of Peptide:N-Glycanase Results in Enha