Open AccessCloning, expression, purification and characterization of replication protein from plasmid pGP2 from Acetobacter estunensis
Author(s) -
Peter Grones,
Jozef Grones
Publication year - 2010
Publication title -
advances in bioscience and biotechnology
Language(s) - English
Resource type - Journals
eISSN - 2156-8502
pISSN - 2156-8456
DOI - 10.4236/abb.2010.15055
Subject(s) - biology , plasmid , cloning (programming) , microbiology and biotechnology , fusion protein , escherichia coli , tandem affinity purification , affinity chromatography , dna , expression vector , recombinant dna , myc tag , dna replication , biochemistry , gene , enzyme , computer science , programming language
The Acetobacter estunensis Rep34 protein participates in the replication of bacterial plasmid pGP2. The Rep34 protein of the A. estunensis, was cloned to the expression vector, that ensure fusion with a His-tag sequence (Rep34 His-tagged), over-expressed in Escherichia coli and purified by metal-affinity chromatography to yield a highly purified and active protein. On this purified protein number different activities and motifs were detected. DNA band-shift assays showed that the Rep34 His-tagged protein bound to the regulation region for replication on the linear double-stranded DNA. In the protein was determined phosphatase activity, ATPase activity and protein is possible to unwind double strand DNA