Cell-free Synthesis of Rat Preproinsulins: Characterization and Partial Amino Acid Sequence Determination

Overview

Journal Proc Natl Acad Sci U S A

Specialty Science

Date 1976 Jun 1

PMID 778852

Citations 62

Authors

S J Chan

P Keim

D F Steiner

Affiliations

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Abstract

Whole nucleic acid fractions of isolated rat islets of Langerhans greatly stimulate incorporation of radioactive amino acids into protein in a wheat germ ribosomal system. Approximately 30% of the synthetic product is precipitated with antisera to insulin or proinsulin. Characterization of this material by gel chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicates a molecular mass of 11,500 daltons. Trypsin digestion releases intact A chain as well as tryptic fragments of the C-peptides and B chains of the two rat proinsulins. Automated sequence determination of labeled cell-free product purified by immunoprecipitation discloses the presence of 23 additional amino acids NH2-terminal to the B chain sequence of proinsulin. The partial amino acid sequence of this extension is as follows: NH2-X-Leu (Lys) Met-x-Phe-Leu-Phe-Leu-Leu (Lys) Leu-Leu-x-leu-X-X-X-X-X-X-X-X-proinsulin. On the basis of the above evidence we have designated this peptide preproinsulin.

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