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PMID: 6093048 Published · ppublish English Journal Article Research Support, U.S. Gov't, Non-P.H.S. Research Support, U.S. Gov't, P.H.S.

HhaI methylase and restriction endonuclease as probes for B to Z DNA conformational changes in d(GCGC) sequences.

Nucleic acids research ·Vol. 12 ·No. 20 ·1984-10-25 ·Pages 7677-92

Zacharias W, Larson JE, Kilpatrick MW, Wells RD

Abstract

The capacity of the modification methylase (MHhaI) and restriction endonuclease (HhaI) form Haemophilus haemolyticus to methylate and cleave, respectively, recognition sites which are in right-handed B or left-handed Z structures was determined in vitro. Plasmids containing tracts of (dC-dG) as well as numerous individual d(GCGC) sites distributed around the vector were studied. Negative supercoiling was used to convert the (dC-dG) tracts (approximately 30 bp in length) from a right-handed to a left-handed conformation. (Methyl-3H)-SAM was used to localize and quantitate modified d(GCGC) recognition sites, whereas cleavage by HhaI was used to detect unmethylated sites. In the left-handed Z-form, the (dC-dG) blocks were not methylated by MHhaI and not cleaved by HhaI. A two-dimensional gel analysis of a family of 33 topoisomers treated with MHhaI revealed that the lack of methylation in the (dC-dG) blocks was directly correlated to the supercoil-induced B to Z transition in these segments. These results are significant with respect to enzyme-DNA interactions in general and provide the basis for using HhaI and MHhaI as probes for different DNA structures and conformational transitions under physiological conditions.

MeSH Terms
Base Sequence DNA DNA (Cytosine-5-)-Methyltransferases/metabolism DNA Restriction Enzymes/metabolism DNA-Cytosine Methylases Deoxyribonucleases, Type II Site-Specific Haemophilus/enzymology Kinetics Methylation Methyltransferases/metabolism Nucleic Acid Conformation Plasmids S-Adenosylmethionine/metabolism
Chemicals
S-Adenosylmethionine DNA DNA modification methylase HhaI DNA-Cytosine Methylases Methyltransferases DNA (Cytosine-5-)-Methyltransferases DNA Restriction Enzymes Deoxyribonucleases, Type II Site-Specific GCGC-specific type II deoxyribonucleases
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Zacharias W
Larson J E
Kilpatrick M W
Wells R D
References (38)
38 references, click to expand
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Article Info
Journal
Nucleic acids research
Abbr.
Nucleic Acids Res
ISSN
0305-1048
DOI
10.1093/nar/12.20.7677
Published
1984-10-25
Pages
7677-92
Language
English
Region
England
NLM ID
0411011
PMCID
PMC320193
Subset
IM
Grants
NIGMS NIH HHS · GM-21839 · United States
External Links
PubMed source DOI: 10.1093/nar/12.20.7677 PMC Full Text View journal details
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