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PMID: 1650347 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Characterization and expression of the Escherichia coli Mrr restriction system.

Journal of bacteriology ·Vol. 173 ·No. 16 ·1991-08-00 ·Pages 5207-19

Waite-Rees PA, Keating CJ, Moran LS, Slatko BE, Hornstra LJ, Benner JS

Abstract

The mrr gene of Escherichia coli K-12 is involved in the acceptance of foreign DNA which is modified. The introduction of plasmids carrying the HincII, HpaI, and TaqI R and M genes is severely restricted in E. coli strains that are Mrr+. A 2-kb EcoRI fragment from the plasmid pBg3 (B. Sain and N. E. Murray, Mol. Gen. Genet. 180:35-46, 1980) was cloned. The resulting plasmid restores Mrr function to mrr strains of E. coli. The boundaries of the mrr gene were determined from an analysis of subclones, and plasmids with a functional mrr gene produce a polypeptide of 33.5 kDa. The nucleotide sequence of the entire fragment was determined; in addition to mrr, it includes two open reading frames, one of which encodes part of the hsdR. By using Southern blot analysis, E. coli RR1 and HB101 were found to lack the region containing mrr. The acceptance of various cloned methylases in E. coli containing the cloned mrr gene was tested. Plasmid constructs containing the AccI, CviRI, HincII, Hinfl (HhaII), HpaI, NlaIII, PstI, and TaqI N6-adenine methylases and SssI and HhaI C5-cytosine methylases were found to be restricted. Plasmid constructs containing 16 other adenine methylases and 12 cytosine methylases were not restricted. No simple consensus sequence causing restriction has been determined. The Mrr protein has been overproduced, an antibody has been prepared, and the expression of mrr under various conditions has been examined. The use of mrr strains of E. coli is suggested for the cloning of N6-adenine and C5-cytosine methyl-containing DNA.

Related Genes
mrr
MeSH Terms
Amino Acid Sequence Base Sequence Blotting, Southern Blotting, Western Cloning, Molecular DNA Restriction Enzymes/biosynthesis,chemistry,genetics DNA-Cytosine Methylases/metabolism Endonucleases/genetics Escherichia coli/genetics,physiology Escherichia coli Proteins Gene Expression/physiology Molecular Sequence Data Open Reading Frames/genetics Site-Specific DNA-Methyltransferase (Adenine-Specific)/metabolism
Chemicals
Escherichia coli Proteins DNA-Cytosine Methylases Site-Specific DNA-Methyltransferase (Adenine-Specific) Endonucleases DNA Restriction Enzymes Mrr protein, E coli
Authors & Affiliations
6 authors, click to expand affiliations / ORCID
Waite-Rees P A
New England Biolabs Inc., Beverly, Massachusetts 01915.
Keating C J
Moran L S
Slatko B E
Hornstra L J
Benner J S
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Article Info
Journal
Journal of bacteriology
Abbr.
J Bacteriol
ISSN
0021-9193
DOI
10.1128/jb.173.16.5207-5219.1991
Published
1991-08-00
Pages
5207-19
Language
English
Region
United States
NLM ID
2985120R
PMCID
PMC208215
Subset
IM
Databases
GENBANK
M64121, M64122, M64123, M64124, M64125, M64126, M64127, M64128, M64129, X54198
External Links
PubMed source DOI: 10.1128/jb.173.16.5207-5219.1991 PMC Full Text View journal details
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