Purification and characterization of angiotensin-binding protein from porcine liver cytosolic fraction.
A protein that binds angiotensins with high affinity was found in porcine liver cytosol, purified to apparent homogeneity and characterized. The protein was named soluble angiotensin-binding protein (sABP) to distinguish it from angiotensin II receptors present on plasma membranes. Purification of the protein was achieved by a combination of ammonium sulfate fractionation, hydrophobic chromatography, ion-exchange chromatography, hydroxylapatite column chromatography and Mono Q ion-exchange chromatography. Specific angiotensin-binding activity, as measured using 125I-angiotensin II, was enriched more than 3400-fold. SDS/polyacrylamide gel electrophoresis of the purified sABP yielded a single 75-kDa protein band, in good agreement with the molecular mass estimated by affinity labeling. sABP was very similar to the angiotensin II receptor in its sensitivity to reducing agents and in its affinities for angiotensin analogues ([Sar1, Ala8]angiotensin II greater than angiotensin III greater than angiotensin II greater than angiotensin I), suggesting a possible similarity between the ligand-binding sites of sABP and the angiotensin II receptor. To obtain a clue to its physiological role(s), we examined the tissue distribution of sABP and found that this protein is widely distributed not only in the peripheral organs but also in the brain.
Hagiwara
H
H
Department of Biological Sciences, Tokyo Institute of Technology, Japan.
Sugiura
N
N
Wakita
K
K
Hirose
S
S
D000345:Affinity Labels; D000804:Angiotensin II / Q000378:metabolism*; D000818:Animals; D001667:Binding, Competitive; D002352:Carrier Proteins / Q000302:isolation & purification* / Q000378:metabolism; D002845:Chromatography / Q000379:methods; D003600:Cytosol / Q000032:analysis*; D004229:Dithiothreitol; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008099:Liver / Q000032:analysis*; D008970:Molecular Weight; D009928:Organ Specificity; D013438:Sulfhydryl Compounds; D013552:Swine
D000345:Affinity Labels; D002352:Carrier Proteins; D013438:Sulfhydryl Compounds; D000804:Angiotensin II; D004229:Dithiothreitol