Skip to main content

DNA determinants important in sequence recognition by Eco RI endonuclease.

Publication ,  Journal Article
Lu, AL; Jack, WE; Modrich, P
Published in: J Biol Chem
December 25, 1981

Alkylation interference and protection methods (Siebenlist, U., and Gilbert, W., (1980) Proc. Natl. Acad. Sci. U. S. A. 77, 122-126) have been utilized to deduce potential DNA contacts involved in specific complex formation between Eco RI endonuclease and its recognition sequence. The endonuclease protected the N7 position (major groove) of the dG and the N3 position (minor groove) of both dA residues within the Eco RI sequence against alkylation by dimethylsulfate, d(GpApApTpTpC), suggesting the presence of poly-peptide in both grooves in the vicinity of affected nitrogens. Results of methylation interference analysis suggest that the N7 of the Eco RI site dG and the N3 of the central dA, d(GpApApTpTpC), are utilized as contacts by the enzyme. The failure to observe interference upon methylation of the 5'-penultimate dA within the sequence implies that the endonuclease does not bond to the N3 of this residue, despite the fact that it is protected against alkylation by the protein. Ethylation interference patterns suggest four major phosphate contacts between endonuclease and each DNA strand. Two of these phosphates are 5'-external to the Eco RI sequence, d(pNpGpApApTpTpC), suggesting involvement of outside phosphates in electrostatic interactions. Moreover, alkylation protection and interference effects on the two DNA strands display perfect 2-fold symmetry. Thus, the endonuclease interacts with a minimum of 10 nucleotide pairs to yield a DNA-protein complex characterized by elements of symmetry. In contrast, specific alkylation effects were not observed in comparable experiments with the endonuclease and a DNA which had been previously methylated by the Eco RI modification enzyme.

Duke Scholars

Published In

J Biol Chem

ISSN

0021-9258

Publication Date

December 25, 1981

Volume

256

Issue

24

Start / End Page

13200 / 13206

Location

United States

Related Subject Headings

  • Substrate Specificity
  • Protein Binding
  • Plasmids
  • Escherichia coli
  • DNA, Bacterial
  • DNA Restriction Enzymes
  • Biochemistry & Molecular Biology
  • Base Sequence
  • 34 Chemical sciences
  • 32 Biomedical and clinical sciences
 

Citation

APA
Chicago
ICMJE
MLA
NLM
Lu, A. L., Jack, W. E., & Modrich, P. (1981). DNA determinants important in sequence recognition by Eco RI endonuclease. J Biol Chem, 256(24), 13200–13206.
Lu, A. L., W. E. Jack, and P. Modrich. “DNA determinants important in sequence recognition by Eco RI endonuclease.J Biol Chem 256, no. 24 (December 25, 1981): 13200–206.
Lu AL, Jack WE, Modrich P. DNA determinants important in sequence recognition by Eco RI endonuclease. J Biol Chem. 1981 Dec 25;256(24):13200–6.
Lu, A. L., et al. “DNA determinants important in sequence recognition by Eco RI endonuclease.J Biol Chem, vol. 256, no. 24, Dec. 1981, pp. 13200–06.
Lu AL, Jack WE, Modrich P. DNA determinants important in sequence recognition by Eco RI endonuclease. J Biol Chem. 1981 Dec 25;256(24):13200–13206.

Published In

J Biol Chem

ISSN

0021-9258

Publication Date

December 25, 1981

Volume

256

Issue

24

Start / End Page

13200 / 13206

Location

United States

Related Subject Headings

  • Substrate Specificity
  • Protein Binding
  • Plasmids
  • Escherichia coli
  • DNA, Bacterial
  • DNA Restriction Enzymes
  • Biochemistry & Molecular Biology
  • Base Sequence
  • 34 Chemical sciences
  • 32 Biomedical and clinical sciences