Glycoproteomic measurement of site-specific polysialylation.

Glycoproteomic measurement of site-specific polysialylation. Anal Biochem. 2020 Feb 20;:113625 Authors: Pelingon R, Pegg CL, Zacchi LF, Phung TK, Howard CB, Xu P, Hardy MP, Owczarek CM, Schulz BL Abstract Polysialylation is the enzymatic addition of a highly negatively charged sialic acid polymer to the non-reducing termini of glycans. Polysialylation plays an important role in development, and is involved in neurological diseases, neural tissue regeneration, and cancer. Polysialic acid (PSA) is also a biodegradable and non-immunogenic conjugate to therapeutic drugs to improve their pharmacokinetics. PSA chains vary in length, composition, and linkages, while the specific sites of polysialylation are important determinants of protein function. However, PSA is difficult to analyse by mass spectrometry (MS) due to its high negative charge and size. Most analytical approaches for analysis of PSA measure its degree of polymerization and monosaccharide composition, but do not address the key questions of site specificity and occupancy. Here, we developed a high-throughput LC-ESI-MS/MS glycoproteomics method to measure site-specific polysialylation of glycoproteins. This method measures site-specific PSA modification by using mild acid hydrolysis to eliminate PSA and sialic acids while leaving the glycan backbone intact, together with protease digestion followed by LC-ESI-MS/MS glycopeptide detection. PSA-modified glycopeptides are not det...
Source: Analytical Biochemistry - Category: Biochemistry Authors: Tags: Anal Biochem Source Type: research