Location of the active site for enzyme-adenylate formation in DNA ligases - PubMed (original) (raw)

Comparative Study

Location of the active site for enzyme-adenylate formation in DNA ligases

A E Tomkinson et al. Proc Natl Acad Sci U S A. 1991.

Abstract

The enzyme-AMP reaction intermediate of the 102-kDa bovine DNA ligase I was digested with trypsin, and the adenylylated peptide was isolated by chromatography under conditions that maintain the acid-labile phosphoramidate bond. Microsequencing of the peptide showed that it contains an internal trypsin-resistant lysine residue, as expected for the site of adenylylation. Inhibition of DNA ligase I activity by pyridoxal 5'-phosphate also indicated the presence of a reactive lysine residue in the catalytic domain of the enzyme. Comparison of the known primary structures of several other DNA ligases with the adenylylated region of mammalian DNA ligase I allows their active sites to be tentatively assigned by sequence homology. The ATP-dependent DNA ligases of mammalian cells, fission yeast, budding yeast, vaccinia virus, and bacteriophages T3, T4, and T7 contain the active site motif Lys-Tyr/Ala-Asp-Gly-(Xaa)-Arg, with the reactive lysine residue flanked by hydrophobic amino acids. The distance between the postulated adenylylation site and the carboxyl terminus of the polypeptide is very similar in these ATP-dependent DNA ligases, whereas the size of the amino-terminal region is highly variable.

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References

    1. Nature. 1970 Aug 15;227(5259):680-5 - PubMed
    1. Biochemistry. 1990 Jul 3;29(26):6132-8 - PubMed
    1. J Biol Chem. 1973 Jan 25;248(2):672-5 - PubMed
    1. J Biol Chem. 1975 Nov 10;250(21):8438-44 - PubMed
    1. Anal Biochem. 1976 May 7;72:248-54 - PubMed

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