Tyrosine 27 of the specificity polypeptide of EcoKI can be UV crosslinked to a bromodeoxyuridine-substituted DNA target sequence

ABSTRACT The specificity (S) subunit of the restriction enzyme EcoKI imparts specificity for the sequence AAC(N6) Substitution of thymine with bromodeoxyuridine in a 25 bp DNA duplex containing this sequence stimulated UV light-induced covalent cross-linking to the S subunit. Crosslinking occurred o...

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Veröffentlicht in:Nucleic acids research 1995-04, Vol.23 (7), p.1177-1183
Hauptverfasser: Chen, A., Powell, L. M., Dryden, D. T. F., Murray, N. E., Brown, T.
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Sprache:eng
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Zusammenfassung:ABSTRACT The specificity (S) subunit of the restriction enzyme EcoKI imparts specificity for the sequence AAC(N6) Substitution of thymine with bromodeoxyuridine in a 25 bp DNA duplex containing this sequence stimulated UV light-induced covalent cross-linking to the S subunit. Crosslinking occurred only at the residue complementary to the first adenine in the AAC sequence, demonstrating a close contact between the major groove at this sequence and the S subunit. Peptide sequencing of a proteolyticallydigested, crosslinked complex identified tyrosine 27 in the S subunit as the site of crosslinking. This is consistent with the role of the N-terminal domain of the S subunit in recognizing the AAC sequence. Tyrosine 27 is conserved in the S subunits of the three type I enzymes that share the sequence AA in the trinucleotide component of their target sequence. This suggests that tyrosine 27 may make a similar DNA contact in these other enzymes.
ISSN:0305-1048
1362-4962
DOI:10.1093/nar/23.7.1177