Location of the carbohydrates present in the HK-ATPase vesicles isolated from hog gastric mucosa

Journal Article · Tue Jan 23 04:00:00 EST 1990 · Biochemistry; (USA)

DOI:https://doi.org/10.1021/bi00455a016· OSTI ID:6747165

Hall, K; Perez, G; Anderson, D; Gutierrez, C; Munson, K; Hersey, S J; Kaplan, J H; Sachs, G ^[1]

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1. CURE, Veterans Administration Medical Center, Los Angeles, CA (USA)

The glycosylation of H+K(+)-ATPase vesicles isolated from hog gastric mucosa was investigated by various methods. Following protein separation on sodium dodecyl sulfate reducing gels and transfer to poly(vinyl difluoride) membranes, binding of concanavalin A was confined to the 94-kDa band which corresponds to the catalytic subunit. In contrast, wheat germ agglutinin binding occurred in a region below the 94-kDa subunit, corresponding to the 60-85-kDa region, and also to protein just above the catalytic subunit. Treatment with glycopeptidase F removed most of the concanavalin A staining and also the wheat germ agglutinin staining found below the 94-kDa region, but spared the higher molecular weight wheat germ agglutinin reactive material. During the deglycosylation experiments a protein of 35-kDa was produced. Sequencing analysis of V8 protease generated peptide fragments of the 35-kDa protein show at least 30% homology with the Na+K(+)-ATPase beta-subunits. Labeling of the carbohydrates by galactosyltransferase and (3H)uridine diphosphate-galactose showed that the sites of labeling were extracellular and were confined to the wheat germ agglutinin staining regions. Two molecular weight regions, below the 94-kDa region, of 60 and 85 kDa were identified. Electron microscopy using postembedding staining techniques showed that both concanavalin A and wheat germ agglutinin staining occurred on the extracellular face of the gastric vesicles.

• OSTI ID: 6747165
• Journal Information: Biochemistry; (USA), Journal Name: Biochemistry; (USA) Vol. 29:3; ISSN 0006-2960; ISSN BICHA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ACID ANHYDRASES
AGGLUTININS
AMINO ACID SEQUENCE
ANIMALS
ANTIBODIES
ATP-ASE
BIOCHEMICAL REACTION KINETICS
BIOLOGICAL LOCALIZATION
CARBOHYDRATES
CONCANAVALIN
DIGESTIVE SYSTEM
DOMESTIC ANIMALS
ELECTRON MICROSCOPY
ELECTROPHORESIS
ENZYMES
GASTROINTESTINAL TRACT
GLYCOPROTEINS
HYDROGEN COMPOUNDS
HYDROLASES
ISOTOPE APPLICATIONS
KINETICS
MAMMALS
MEMBRANES
MICROSCOPY
MOLECULAR STRUCTURE
MOLECULAR WEIGHT
MUCOUS MEMBRANES
ORGANIC COMPOUNDS
PHOSPHOHYDROLASES
PROTEINS
REACTION KINETICS
SWINE
TRACER TECHNIQUES
TRITIUM COMPOUNDS
VERTEBRATES