Production and characterization of a monoclonal antibody to H+ATPase

Conference · Sat Mar 01 04:00:00 EST 1986 · Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States)

OSTI ID:5278211

Yurko, M; Fitch, F; Gluck, S

Acidification of endocytic vesicles is carried out by an ATP-dependent proton pump, H+ATPase, an FOF1 type enzyme comprised of at least 5 major subunits of 70, 56, 45, 35, and 17 kDa. A monoclonal antibody, H6.1, to H+ATPase from bovine kidney medulla, was raised to enable the structural characterization and localization of the pump. Several criteria were used to show that H6.1 recognized H+ATPase. 1.) H6.1 immunoprecipitated N-ethylmaleimide-sensitive and vanadate- and azide-insensitive solubilized ATPase activity (and GTPase activity) from both crude and purified enzyme preparations. 2.) H6.1 immunoprecipitated oligomycin-insensitive ATP-dependent proton transporting vesicles made from bovine kidney medulla, rat kidney, and CHO cells. 3.) H6.1 specifically immuno-precipitated the 5 subunits of H+ATPase from a partially purified preparation of the enzyme that had been labelled with I-125. H6.1 was then used as an immunocytochemical probe for the localization of H+ATPase. In bovine kidney medullary collecting duct, there was an intense apical staining of selected cells. In proximal tubule and in cultured CHO cells there was a granular pattern of staining characteristic of endocytic vesicles and lysosomes, suggesting that the kidney and CHO cell proton pumps are structurally related.

• Research Organization: Univ. of Chicago, IL
• OSTI ID: 5278211
• Report Number(s): CONF-8604222-
• Conference Information: Journal Name: Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States) Journal Volume: 45:3
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ACID ANHYDRASES
ANIMAL CELLS
ANIMALS
ANTIBODIES
ATP-ASE
BETA DECAY RADIOISOTOPES
BIOLOGICAL LOCALIZATION
BIOSYNTHESIS
BODY
CATTLE
CHEMICAL COMPOSITION
CHO CELLS
DAYS LIVING RADIOISOTOPES
DOMESTIC ANIMALS
ELECTRON CAPTURE RADIOISOTOPES
ENZYME ACTIVITY
ENZYMES
HYDROLASES
INTERMEDIATE MASS NUCLEI
IODINE 125
IODINE ISOTOPES
ISOTOPE APPLICATIONS
ISOTOPES
KIDNEYS
MAMMALS
MEMBRANE TRANSPORT
MONOCLONAL ANTIBODIES
NUCLEI
ODD-EVEN NUCLEI
ORGANS
PHOSPHOHYDROLASES
RADIOISOTOPES
RATS
RODENTS
RUMINANTS
SYNTHESIS
TRACER TECHNIQUES
VERTEBRATES