Amino-terminal amino acid sequence and heterogeneity in glycosylation of rat renal renin.
We isolated 7.4 mg of pure renin from 2 kg of rat kidneys using affinity chromatography on pepstatin-aminohexyl-Sepharose and an octapeptide renin inhibitor, H-77-Sepharose. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that renin consists of two polypeptide chains linked by a disulfide bond, one of Mr = 36,000 (heavy chain) and the other of Mr = 3,000 (light chain). The amino-terminal 10-amino acid sequences of the heavy and the light chains were identical to the sequences beginning at Ser72 and Asp355, respectively, of the amino acid sequence of preprorenin deduced from the renin cDNA sequence. Amino acid sequencing of the carboxyl-terminal peptide of the heavy chain, generated by digestion with lysyl endopeptidase, showed that the carboxyl-terminal residue of the heavy chain is Phe. Thus, the propeptide of prorenin is cleaved after Thr71, followed by removal of two amino acids, Arg353 and Asn354, the result being formation of the heavy and light chains. Thus, the site of cleavage of rat prorenin is after a nonbasic amino acid, in contrast to the cleavage of the propeptide after a pair of basic amino acids in mouse submaxillary renin, human renal renin, and many secretory proteins. Treatment of renin with neuraminidase or glycopeptidase F had no apparent effect on the charge heterogeneity of renin. Glycosylation probably does not contribute to charge heterogeneity.
Kim
S
S
Department of Pharmacology, Osaka City University Medical School, Japan.
Hosoi
M
M
Kikuchi
N
N
Yamamoto
K
K
D000595:Amino Acid Sequence; D000818:Animals; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D004792:Enzyme Precursors / Q000235:genetics; D006031:Glycosylation; D007668:Kidney / Q000201:enzymology*; D007700:Kinetics; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010449:Peptide Mapping; D011499:Protein Processing, Post-Translational; D051381:Rats; D012083:Renin / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism
D004792:Enzyme Precursors; D046911:Macromolecular Substances; D012083:Renin