Release of Oligosaccharides from Human Erythrocyte Membranes of Different Blood-Group-P Phenotypes by Trifluoroacetolysis

Abstract

Human erythrocyte membranes of different blood-group-P phenotypes have been degraded by trifluoroacct olysis at conditions that will effect transamidation. This degradation resulted in dissolution of the membranes with concomitant degradation of proteins. Oligosaccharide chains linked to proteins and ceramides were released as their pertrifluoroacetylated derivatives, which generally are stable towards further action of the reagent. 2-Acetamido-2-deoxy-sugar residues were converted into their corresponding 2-deoxy-2-trifluoroacetamido derivatives. The mixture of pertrifluoroacetylated oligosaccharides obtained was de-0-trifluoroacetylated and de-N-trifluoroacetylated, reduced, acetylated, permethylated and analyzed by gas-liquid chromatography/mass spectrometry. Membranes obtained from two individuals with phenotype B, p (known to contain an excess of lactosyl ceramide) gave almost identical results with lactose released as the main component. From AZB,Pf cell membranes small amounts of lactose were obtained but the trisaccharide Gal(al-4)Gal(~1-4)Glc was the major component found. This trisaccharide is most likely derived from the Pk antigen (trihexosyl ceramide). Membranes from a donor with the phenotype B,P1 yielded small amounts of both lactose and the Pk trisaccharide together with a larger quantity of the tetrasaccharide GalNAc(~l-3)Gal(al-4)Gal(/J1-4)Glc. The tetrasaccharide represents the carbohydrate portion of the P antigen (globoside). From all three types of membranes the B-specific trisaccharide Gal(a1-3)Gal I ( a 1 4 Fuc a;i\ obtained. This compound is probably derived from specific degradation of blood-group-B-active glycoproteins and/or glycolipids. In addition to the oligosaccharides mentioned above a large number of others were observed but in lower amounts.Recently a new reaction, trifluoroacetolysis, has been developed which will effect transamidation at conditions where most glycosides and reducing sugars are stable, being converted into their pertrifluoroacetylated derivatives [I -41. Using this reaction procedures for the isolation of 0-and N-glycosidically linked carbohydrate chains in glycoproteins have been developed [5-71. It has also been demonstrated that the glycosidic bond to the sphingosine moiety in glycolipids is cleaved specifically by trifluoroacetolysis [8].Cell surface carbohydrates exist in the form of monosaccharides and oligosaccharides linked either to proteins or lipids which are buried in the cell mem-

Characterization of a UDP-Gal:Galβ1–3GalNAc α1,4-Galactosyltransferase Activity in a Mamestra brassicaeCell Line

“…Interestingly, chemical and immunochemical studies have indicated that galabiosylceramide and P 1 and P k antigens are expressed in many pig and human tissues (73,(77)(78)(79). In addition, several studies have shown that Gal␣1-4Gal-containing glycolipids act as receptors for the binding of pathogenic bacteria, such as Escherichia coli to human uroepithelial cells (79) and Streptococcus suis, which is responsible for meningitis in pigs and humans (80,81).…”

Glycolipid Analysis

“…39 O-trifluoroacetylation is complete and amino functions are all N-trifluoroacetylated. N-Acetyl groups undergo transamidation to N-trifluoroacetyl groups but sialic acid residues are hydrolyzed from the gangliosides.…”