In vitro translation of avian vitellogenin messenger RNA

J. I. Gordon, R. G. Deeley, A. T.H. Burns, B. M. Paterson, J. L. Christmann, R. F. Goldberger

Research output: Contribution to journalArticlepeer-review

26 Scopus citations

Abstract

Administration of 17β-estradiol to roosters induces the synthesis of vitellogenin in the liver. The mRNA that specifies this protein has been purified from the livers of estrogen-treated roosters and has been shown to have a molecular weight of 2.3 x 106. In order to rigorously establish the identity of the polypeptide specified by this mRNA, we used a staphylococcal nuclease-treated, mRNA-dependent wheat germ cell-free translation system capable of synthesizing polypeptides as large as vitellogenin (monomer M(r) = 240,000). Vitellogenin mRNA directs the in vitro synthesis of a polypeptide with the following features: (a) it co-migrates with authentic vitellogenin in SDS-polyacrylamide gels; (b) it is highly enriched for serine but is not phosphorylated; (c) it is immunoprecipitated by purified, monospecific, anti-vitellogenin antibody; and (d) it has an unusual cyanogen bromide cleavage pattern characteristic of vitellogenin. The most striking characteristic of the cyanogen bromide cleavage products is an extremely large polypeptide (M(r) = 90,000) that contains two phosvitins. The kinetics of incorporation of serine and methionine into vitellogenin synthesized in the wheat germ cell-free translation system indicates that the phosvitins are located near the COOH-terminal portion of the molecule.

Original languageEnglish
Pages (from-to)8320-8327
Number of pages8
JournalJournal of Biological Chemistry
Volume252
Issue number22
StatePublished - 1977

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