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Removal of a frameshift between the hsdM and hsdS genes of the EcoKI Type IA DNA restriction and modification system produces a new type of system and links the different families of Type I systems.

TitleRemoval of a frameshift between the hsdM and hsdS genes of the EcoKI Type IA DNA restriction and modification system produces a new type of system and links the different families of Type I systems.
Publication TypeJournal Article
Year of Publication2012
AuthorsRoberts GA, Chen K, Cooper LP, White JH, Blakely GW, Dryden DTF
JournalNucleic Acids Res
Volume40
Issue21
Pagination10916-24
Date Published2012 Nov
ISSN1362-4962
KeywordsArtificial Gene Fusion, Bacterial Proteins, Coliphages, Deoxyribonucleases, Type I Site-Specific, DNA Cleavage, DNA Restriction Enzymes, DNA Restriction-Modification Enzymes, Escherichia coli, Escherichia coli Proteins, Frameshifting, Ribosomal, Mutagenesis, Protein Subunits, Recombinant Fusion Proteins, Site-Specific DNA-Methyltransferase (Adenine-Specific), Transformation, Bacterial
Abstract

The EcoKI DNA methyltransferase is a trimeric protein comprised of two modification subunits (M) and one sequence specificity subunit (S). This enzyme forms the core of the EcoKI restriction/modification (RM) enzyme. The 3' end of the gene encoding the M subunit overlaps by 1 nt the start of the gene for the S subunit. Translation from the two different open reading frames is translationally coupled. Mutagenesis to remove the frameshift and fuse the two subunits together produces a functional RM enzyme in vivo with the same properties as the natural EcoKI system. The fusion protein can be purified and forms an active restriction enzyme upon addition of restriction subunits and of additional M subunit. The Type I RM systems are grouped into families, IA to IE, defined by complementation, hybridization and sequence similarity. The fusion protein forms an evolutionary intermediate form lying between the Type IA family of RM enzymes and the Type IB family of RM enzymes which have the frameshift located at a different part of the gene sequence.

DOI10.1093/nar/gks876
Alternate JournalNucleic Acids Res.
PubMed ID23002145
PubMed Central IDPMC3510504
Grant List090288/Z/09/ZA / / Wellcome Trust / United Kingdom
GR080463MA / / Wellcome Trust / United Kingdom
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