title: Development of a method for the efficient release of N-glycans from glycoproteins generating native deglycosylated proteins.
abstract: In many cases, it is desirable to maintain the native status of the target glycoproteins when they are deglycosylated. However, most conventional deglycosylation process often causes the irreversible denaturation of the target glycoproteins. In the present study, we developed a deglycosylation method that could obtain the native deglycosylated proteins employing Png1p-ΔH1, which was confirmed to tolerate high concentration of dithiothreitol (DTT). To prove this process, ribonuclease B (RNase B) and Yeast carboxypeptidase (CPY) were employed as the targeting glycoproteins. Our results confirmed that both of them could be completely deglycosylated in the presence of high concentration DTT and could be refolded when DTT was removed. The circular dichroism spectroscopy (CD) measurement of refolded CPY and RNase B indicated that the structure of deglycosylated proteins had recovered their native status. This method offers the possibility of efficiently releasing N-linked glycans from glycoproteins and obtaining the native target proteins.
authors: State Key Laboratory of Microbial Technology, Shandong University, Jinan, Shandong 250100, PR China.
authors: Shengjun
authors: S
authors: Wang
authors: Chuanliang
authors: C
authors: Ma
authors: Huili
authors: H
authors: Yu
authors: Changgan
authors: C
authors: Huang
authors: Qingsheng
authors: Q
authors: Qi
mesh_terms: D001709:Biotechnology / Q000379:methods*; D002268:Carboxypeptidases / Q000737:chemistry / Q000378:metabolism*; D002942:Circular Dichroism; D004229:Dithiothreitol; D006023:Glycoproteins / Q000737:chemistry / Q000378:metabolism*; D006031:Glycosylation; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase / Q000235:genetics / Q000378:metabolism*; D011134:Polysaccharides / Q000378:metabolism*; D011489:Protein Denaturation; D058849:Protein Refolding; D012260:Ribonucleases / Q000737:chemistry / Q000378:metabolism*
chemical_list: D006023:Glycoproteins; D011134:Polysaccharides; D012260:Ribonucleases; C029600:ribonuclease B; D002268:Carboxypeptidases; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; D004229:Dithiothreitol