A purification method and N-glycosylation sites of a 36-cysteine-containing, putative cell/cell adhesion glycoprotein gp64 of the cellular slime mold, Polysphondylium pallidum.
A 64-kDa membrane-bound glycoprotein (gp64) of the cellular slime mold Polysphondylium pallidum, is a putative cell/cell adhesion protein identified by adhesion-blocking antibody fragments (Fab). gp64 is expressed on the cell surface of growth-phase cells and seems to mediate cell/cell adhesion. This paper describes an improved purification method based on the lipophilic nature of this protein. A critical step in the purification method is to collect an insoluble top layer appearing during ammonium sulfate precipitation. The sequence of cDNA encoding gp64 and its deduced amino acid sequence have been determined previously. Based on cDNA sequence data, the structure of gp64 protein was analyzed: almost all amino acid compositions and partial amino acid sequences of lysylendopeptidase-digested peptides of gp64 were determined by protein analysis; all six asparagine-linked glycosylation sites (Asn-Xaa-Ser/Thr) in fact contain carbohydrates, and all 36 cysteine residues were involved in forming disulfide bridges. From these data, gp64 seems to be a unique protein among cell/cell adhesion proteins.
Saito
T
T
Department of Botany, Faculty of Science, Hokkaido University, Sapporo, Japan.
Kumazaki
T
T
Ochiai
H
H
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D015815:Cell Adhesion Molecules / Q000302:isolation & purification* / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D003545:Cysteine / Q000737:chemistry; D004220:Disulfides / Q000032:analysis; D006020:Glycopeptides / Q000737:chemistry; D006023:Glycoproteins / Q000302:isolation & purification* / Q000378:metabolism; D006031:Glycosylation; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009235:Myxomycetes / Q000737:chemistry*; D010446:Peptide Fragments / Q000737:chemistry
D000596:Amino Acids; D015815:Cell Adhesion Molecules; D004220:Disulfides; D006020:Glycopeptides; D006023:Glycoproteins; D010446:Peptide Fragments; D003545:Cysteine