The use of complementary peptides in the purification of an angiotensin II binding protein.
We used the molecular recognition hypothesis, that peptide ligands and their receptor binding sites are encoded by complementary nucleotide sequences, to purify an angiotensin II (Ang II) binding protein. The complementary peptide IIA (Lys-Gly-Val-Asp-Val-Tyr-Ala-Val) specified by the RNA sequence complementary to the messenger (m)RNA sequence for rat Ang II was synthesized, purified and used to raise polyclonal antibodies. Complementary peptide IIA specifically inhibited the binding of 125I-Ang II to receptors on rat adrenal membranes, and anti-IIA immunoglobulin G (IgG) specifically inhibited the binding of 125I-Ang II to rat adrenal Ang II receptors and Ang II-dependent aldosterone secretion by cultured rat adrenal cells, suggesting that the antibody recognizes the Ang II receptor. Anti-IIA IgG was used for immuno-affinity purification, from a rat adrenal membrane preparation of an Ang II binding protein with a molecular weight of 66,000 +/- 2000 that bound 125I-Ang II specifically. This is the first report of purification of an Ang II receptor binding protein which retains its capacity to specifically bind 125I-Ang II.
Elton
T S
TS
Department of Medicine, University of Alabama, Birmingham 35294.
Oparil
S
S
Blalock
J E
JE
D000311:Adrenal Glands / Q000032:analysis*; D000450:Aldosterone / Q000096:biosynthesis; D000804:Angiotensin II / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001667:Binding, Competitive; D002352:Carrier Proteins / Q000302:isolation & purification*; D009842:Oligopeptides / Q000378:metabolism; D051381:Rats; D013329:Structure-Activity Relationship
D002352:Carrier Proteins; D009842:Oligopeptides; D000804:Angiotensin II; D000450:Aldosterone