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Eukaryotic mRNA Capping Enzyme-Guanylate Covalent Intermediate

Sundararajan Venkatesan and Bernard Moss
Proceedings of the National Academy of Sciences of the United States of America
Vol. 79, No. 2, [Part 1: Biological Sciences] (Jan. 15, 1982), pp. 340-344
Stable URL: http://www.jstor.org/stable/12070
Page Count: 5
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Since scans are not currently available to screen readers, please contact JSTOR User Support for access. We'll provide a PDF copy for your screen reader.
Eukaryotic mRNA Capping Enzyme-Guanylate Covalent Intermediate
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Abstract

Incubation of HeLa cell mRNA guanylyltransferase (GTP:mRNA guanylyltransferase, EC 2.7.7.50) with [α -32P]GTP and a divalent cation in the absence of an RNA acceptor results in the formation of a covalent enzyme-guanylate complex. The complex, after purification by phosphocellulose chromatography, can transfer its bound GMP moiety to pyrophosphate, regenerating GTP, or to the 5′-diphosphate end of poly(A), forming a cap structure G(5′)pppA(pA)n. The GMP-polypeptide has a molecular weight of 65,000 and is stable to heating in the presence of sodium dodecyl sulfate. On the basis of the alkali-stable and acid-labile nature of the bond and its susceptibility to nucleophilic attack by hydroxylamine at low pH, the GMP-polypeptide linkage appears to be a phosphoamide bond. After digestion with trypsin, a single GMP-peptide was resolved by two-dimensional electrophoresis and chromatography.

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