Evidence for a Novel O-Linked Sialylated Trisaccharide on Ser-248 of Human Plasminogen 2

Abstract

Human plasminogen, the inactive precursor of plasmin, exists in two major glycoforms. Plasminogen 1 contains an N-linked oligosaccharide at Asn-289 and an Olinked oligosaccharide at Thr-345. Plasminogen 2 is known to contain only an O-linked oligosaccharide at Thr-345. However, plasminogen 2 displays a further well documented microheterogeneity dependent on the Nacetylneuraminic acid content, which has functional consequences with regard to activation of plasminogen. The proposed structure and number of known oligosaccharide linkages in plasminogen 2 is insufficient to account for this microheterogeneity. In the present study, a combination of trypsin digestion, lectin affinity chromatography, Edman degradation amino acid sequence analysis, carbohydrate composition analysis, and mass spectrometry revealed the existence of a novel site for O-linked glycosylation on plasminogen 2 at Ser-248. Direct evidence for the structure of the carbohydrate was obtained from a combination of lectin affinity chromatography, desialylation experiments, and mass spectrometry analysis. These findings provide a structural basis for some of the observed microheterogeneity, and have implications with regard to the known functional consequences of the extent of sialylation of plasminogen.

The Glycosylation and Structure of Human Serum IgA1, Fab, and Fc Regions and the Role of N-Glycosylation on Fcα Receptor Interactions

“…Thr 225 and Thr 236 produced depressed signals (30 and 60% relative to the expected signal for threonine), while hydroxyamino acids at other positions gave the expected yield of an unmodified residue. Depressed yields of hydroxyamino acids are consistent with O-glycosylation, since glycosylated anilinothiozoline-amino acids are not extracted from the sequencer reaction chamber under the conditions used (51)(52)(53).…”

Structure of the Complex of Maclura pomiferaAgglutinin and the T-antigen Disaccharide, Galβ1,3GalNAc

“…For the Gal, several of its hydroxyls are exposed and apparently could be substituted without impairing binding to MPA. There have been reports that jacalin can accept the 2,3Ј-sialylated glycopeptide (26). The natural ligand for the Moraceae lectins is of course unknown, but the binding site should be capable of accepting a polysaccharide with a Gal␤1,3GalNAc␣ internal element as readily as it does the short O-linked glycopeptides.…”

Biophysical Mechanisms Mediating Fibrin Fiber Lysis

“…Type II only has the O-linked T346 glycan and comprises 60% of circulating material. Subsequent studies suggested that some Type II plasminogen could contain a second O-linked glycan at S248 [52] or S339 [53]. Glycosylation plays a dramatic role in regulating the binding of plasmin(ogen) to particular cell types [54] and seems to play a minor role in the activation of plasminogen on fibrin [55].…”