Exploring the DNA mimicry of the Ocr protein of phage T7.
DNA mimic proteins have evolved to control DNA-binding proteins by competing with the target DNA for binding to the protein. The Ocr protein of bacteriophage T7 is the most studied DNA mimic and functions to block the DNA-binding groove of Type I DNA restriction/modification enzymes. This binding prevents the enzyme from cleaving invading phage DNA. Each 116 amino acid monomer of the Ocr dimer has an unusual amino acid composition with 34 negatively charged side chains but only 6 positively charged side chains. Extensive mutagenesis of the charges of Ocr revealed a regression of Ocr activity from wild-type activity to partial activity then to variants inactive in antirestriction but deleterious for cell viability and lastly to totally inactive variants with no deleterious effect on cell viability. Throughout the mutagenesis the Ocr mutant proteins retained their folding. Our results show that the extreme bias in charged amino acids is not necessary for antirestriction activity but that less charged variants can affect cell viability by leading to restriction proficient but modification deficient cell phenotypes.
EastChem School of Chemistry, The University of Edinburgh, The King's Buildings, Edinburgh, EH9 3JZ, UK.
Gareth A
GA
Roberts
Augoustinos S
AS
Stephanou
Nisha
N
Kanwar
Angela
A
Dawson
Laurie P
LP
Cooper
Kai
K
Chen
Margaret
M
Nutley
Alan
A
Cooper
Garry W
GW
Blakely
David T F
DT
Dryden
D002151:Calorimetry; D004247:DNA / Q000737:chemistry; D053837:DNA Cleavage; D004262:DNA Restriction Enzymes / Q000378:metabolism; D004926:Escherichia coli / Q000166:cytology / Q000235:genetics / Q000254:growth & development; D008958:Models, Molecular; D018716:Molecular Mimicry; D009154:Mutation; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific) / Q000378:metabolism; D014764:Viral Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
C440255:Ocr protein, bacteriophage T7; D014764:Viral Proteins; D004247:DNA; C093244:DNA modification methylase EcoKI; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific); D004262:DNA Restriction Enzymes; C089629:endodeoxyribonuclease EcoKI