Solubilization and separation of the human erythrocyte D-glucose transporter covalently and noncovalently photoaffinity-labeled with (/sup 3/H)cytochalasin B

Journal Article · Wed Jan 01 04:00:00 EST 1986 · Proc. Natl. Acad. Sci. U.S.A.; (United States)

DOI:https://doi.org/10.1073/pnas.83.2.479· OSTI ID:6352273

Kurokawa, T; Tillotson, L G; Chen, C C; Isselbacher, K J

The D-glucose transporter in the human erythrocyte membranes was photoaffinity-labeled with (/sup 3/H)cytochalasin B and solubilized with n-octyl ..beta..-D-glucopyranoside (octyl glucoside). (/sup 3/H)Cytochalasin B-bound proteins were further isolated by using Sephadex G-50 chromatography. The amount of (/sup 3/H)cytochalasin B associated with the membrane proteins was approximately 10% of the total radioactivity in the octyl glucoside extract. The solubilized photoaffinity-labeled D-glucose transporter was isolated and found to consist of two major peaks by DEAE-Sephacel chromatography. The radioactivity of peak II was considerably greater than that of peak I. The incorporation of (/sup 3/H)cytochalasin B into both peaks was blocked by the presence of D-glucose during photolysis. These results indicate the (/sup 3/H)cytochalasin B was covalently bound to the D-glucose transporter only in peak II and that peak II could be generated by the photoaffinity labeling of peak I. However, the D-glucose transport activity was associated only with peak I. These findings suggest that the anionic domain of the D-glucose transporter becomes exposed because of the conformational changes of the protein as a result of covalent binding with (/sup 3/H)cytochalasin B by photoaffinity labeling.

• Research Organization: Massachusetts General Hospital, Boston
• OSTI ID: 6352273
• Journal Information: Proc. Natl. Acad. Sci. U.S.A.; (United States), Journal Name: Proc. Natl. Acad. Sci. U.S.A.; (United States) Vol. 83:2; ISSN PNASA
• Country of Publication: United States
• Language: English

Related Subjects

550601* -- Medicine-- Unsealed Radionuclides in Diagnostics
62 RADIOLOGY AND NUCLEAR MEDICINE
ALDEHYDES
BIOCHEMISTRY
BIOLOGICAL MATERIALS
BLOOD
BLOOD CELLS
BODY FLUIDS
CARBOHYDRATES
CELL CONSTITUENTS
CELL MEMBRANES
CHEMICAL BONDS
CHEMISTRY
CHROMATOGRAPHY
CONFORMATIONAL CHANGES
CYTOCHEMISTRY
ENZYME INHIBITORS
ERYTHROCYTES
GLUCOSE
HEXOSES
INHIBITION
ION EXCHANGE CHROMATOGRAPHY
ISOTOPE APPLICATIONS
LABELLED COMPOUNDS
MATERIALS
MEMBRANE PROTEINS
MEMBRANE TRANSPORT
MEMBRANES
MONOSACCHARIDES
ORGANIC COMPOUNDS
PROTEINS
RADIORECEPTOR ASSAY
RECEPTORS
SACCHARIDES
SEPARATION PROCESSES
TRACER TECHNIQUES
TRITIUM COMPOUNDS