Studies on Glycopeptides Derived from Acidic Glycoproteins of Guinea-Pig Serum

Abstract

1. Fraction I, a fraction containing acidic glycoproteins, isolated from guinea-pig serum, was digested with Pronase after removal of sialic acid and a major and a minor glycopeptide fraction were isolated by chromatography with Sephadex G-25 and G-50. 2. The major fraction was examined by various methods and shown to contain several glycopeptides. Estimates of molecular weight of the glycopeptide fractions were obtained. Although some variation appeared to occur, the glycopeptides were not grossly heterogeneous with respect to size. An average prosthetic group was estimated to contain about 15 sugar residues. 3. Aspartic acid was the principal amino acid present in the fractions and in all subfractions of the major fraction investigated. Where examined, ammonia was liberated on acid hydrolysis in approximately equimolar amounts to the aspartic acid present. The carbohydrate composition of the fractions was also determined. 4. The glycopeptides showed relatively little degradation in alkaline solution. 5. These results suggest that an N-acylglycosylamine bond involving aspartic acid forms the major type of linkage between carbohydrate and polypeptide. The isolation of a compound with the composition and chromatographic properties of 2-acetamido-1-(l-beta-aspartamido)-1,2-dideoxy-beta-d-glucose supports this view, and indicates that N-acetylglucosamine is the sugar involved in at least many linkages. 6. Fraction I contains some glycoproteins that are susceptible to Pronase and one or more others that resist digestion before the removal of sialic acid. A brief examination revealed some similarities between prosthetic groups derived from both kinds of glycoprotein.

Studies on the site of biosynthesis of acidic glycoproteins of guinea-pig serum

“…at 850. After dialysis against water, the non-diffusible material was freeze-dried and digested with Pronase as described by Cunningham & Simkin (1966), except that the amount of Pronase added was increased by one-third. The digest was fractionated on a column (81 cm.…”

“…Previous papers have described work on the characteristics and structure of components of an ac-globulin fraction containing acidic glycoproteins, fraction I, isolated from guinea-pig serum (Simkin, Seshadri & Skinner, 1964a; Simkin, Skinner & Seshadri, 1964b;Cunningham & Simkin, 1966).…”

“…Athineos, Thornton & Winzler, 1962;Bergmann, Levine & Spiro, 1962;Barker & Whitehead, 1963). Glucosamine was used to study formation of the carbohydrate groups because N-acetylglucosamine residues participate in linkage with polypeptide in components of fraction I (Cunningham & Simkin, 1966) and also occur in more peripheral positions in the groups (Simkin et al 1964a). In addition, studies in the rat had shown that glucosamine is used efficiently for synthesis of serum glycoproteins, with label found almost exclusively in hexosamine and sialic acid (e.g.…”

Glycoprotein in Cultured Human Skin Fibroblasts

“…Yamashina (12) reported glycoprotein with an N-glycoside bond in ovoalbumin of eggs, in ovomucoid (13), human lXI-acid glycoprotein (14), y-globulin (15), thyroglobulin (16), RNAase B of calf pancreas (17), human transferrin (18), haptoglobin (19), takaamylase A (20), ceruloplasmin (21), and bovine aorta (22). Structural glycoprotein with O-glycoside bond has been found in the human submandibular gland (23,24), guinea pig serum (25) and myeloma globulin. In this latter group of glycoproteins the hydroxy group on serine and threonine conjugates with sugar, consisting mostly of N-acetyl hexasamine.…”

Isolation and fractionation of glycopeptides from porcine thyroglobulin

“…Molecular-weight determination8 of glycopeptide8 by gel filtration. Molecular weights of glycopeptides were determined by gel filtration (Cunningham & Simkin, 1966; in a similar manner and subjected to gel filtration on a Bhatti & Clamp, 1968). Sephadex G-50 (super fine grade) column (1.6 cmx 90cm) of Sephadex G-50 and eluted with equilibrated with O.lM-NaCl was packed into a column water (Fig.…”