摘要:
The present disclosure provides nuclear magnetic resonance (NMR) methods for characterizing mixtures of N-linked glycans. Without limitation, methods of the present disclosure may be useful in characterizing monosaccharide composition, branching, fucosylation, sulfation, phosphorylation, sialylation linkages, presence of impurities and/or efficiency of a labeling procedure (e.g., labeling with a fluorophore such as 2-AB). In certain embodiments, the methods can be used quantitatively. In certain embodiments, the methods can be combined with enzymatic digestion to further characterize glycan mixtures.
摘要:
The present disclosure provides a method for determining the relative conformations of a protein provided in different protein preparations, comprising steps of: (i) obtaining a first 2D NOESY NMR spectrum of a first protein preparation; (ii) obtaining a second 2D NOESY NMR spectrum of a second protein preparation; and (iii); determining whether a protein has a different conformation in the first and second protein preparations by comparing one or more cross-peaks in the first 2D NOESY NMR spectrum with one or more corresponding cross-peaks in the second 2D NOESY NMR spectrum.
摘要:
The present disclosure provides a method for determining the relative conformations of a protein provided in different protein preparations, comprising steps of: (i) obtaining a first 2D NOESY NMR spectrum of a first protein preparation; (ii) obtaining a second 2D NOESY NMR spectrum of a second protein preparation; and (iii); determining whether a protein has a different conformation in the first and second protein preparations by comparing one or more cross-peaks in the first 2D NOESY NMR spectrum with one or more corresponding cross-peaks in the second 2D NOESY NMR spectrum.
摘要:
The present invention provides methods of evaluating a glycoprotein preparation for the absence, presence or amount of an N-acetylhexosamine glycan, e.g., an N-acetylglucosamine glycan.
摘要:
The present disclosure provides methods for analyzing structure and/or composition of N-glycans. Such methods often involve digestion of N-glycans with multiple exoglycosidases. In some embodiments, N-glycans are digested with multiple exoglycosidases simultaneously. In some embodiments, N-glycans are digested with multiple exoglycosidases sequentially. In some embodiments, methods in accordance with the present disclosure involve comparison of cleavage products of N-glycans that have been digested with multiple exoglycosidases simultaneously to N-glycans that have been digested with multiple exoglycosidases sequentially.
摘要:
The present disclosure provides methods for assessing the glycosylation of a target glycoprotein produced by a cell through analysis of cell-surface glycans on the cell. The present disclosure therefore teaches that glycosylation of cell surface proteins can serve as a proxy for glycosylation of other proteins.
摘要:
The present disclosure provides methods of making isotopically-labeled glycans or a mixture of glycans (e.g., a glycan preparation) via reductive amination. Such methods are useful, for example, in the determination of the relative amounts of a glycan species present in two or more glycoprotein preparations using mass spectroscopy. In one aspect, the present disclosure provides a method of making an isotopically-labeled glycan by a glycan with an amine and an isotopically-labeled reducing agent to provide an isotopically-labeled aminated glycan. In certain embodiments, the amine is an isotopically-labeled amine.
摘要:
The present disclosure provides methods for assessing the glycosylation of a target glycoprotein produced by a cell through analysis of cell-surface glycans on the cell. The present disclosure therefore teaches that glycosylation of cell surface proteins can serve as a proxy for glycosylation of other proteins.