The small heat shock protein IbpB of Escherichia coli can accelerate protein disaggregation from inclusion body by Hsp100-Hsp70 re-activation system in vitro. It was therefore hypothesized that overexpression of IbpB might be able to promote protein disaggregation from inclusion body, by which more soluble recombinant proteins would be obtained. The overexpression of IbpB actually enhanced prod...

Expression strains of Escherichia coli BL21(DE3) overproducing the E. coli m(5)C McrA restriction protein were produced by cloning the mcrA coding sequence behind a T7 promoter. The recombinant mcrA minus BL21(DE3) host produces active McrA as evidenced by its acquired ability to selectively restrict the growth of T7 phage containing DNA methylated in vitro by HpaII methylase. The mcrA coding r...

The sigma factor sigma 73 of the obligate intracytoplasmic bacterium Rickettsia prowazekii was overexpressed and purified from Escherichia coli. The rickettsial rpoD gene encoding sigma 73 was cloned into a Ndel-BamHI-cleaved pET-15b vector under control of T7 transcription and translation signals. The recombinant plasmid encoded a 75 kDa fusion protein that was overproduced in E. coli BL21(DE3...

CaCl2 treatment followed by heat shock is the most common method for artificial transformation. Here, the cells were transformed using CaCl2 treatment either with heat shock (standard protocol) or without heat shock (lab protocol) to comprehend the difference in transformation efficiency. The BL21 strain of Escherichia coli (E. coli) was being susceptible using CaCl2 treatment. Some Cells were ...

An Ambler class A β-lactamase gene, bla(CIA-1), was cloned from the reference strain Chryseobacterium indologenes ATCC 29897 and expressed in Escherichia coli BL21. The bla(CIA-1) gene encodes a novel extended-spectrum β-lactamase (ESBL) that shared 68% and 60% identities with the CGA-1 and CME-1 β-lactamases, respectively. bla(CIA-1)-like genes were detected from clinical isolates. In addition...

To isolate novel and useful microbial enzymes from uncultured gastrointestinal microorganisms, a fecal microbial metagenomic library of the pygmy loris was constructed. The library was screened for amylolytic activity, and 8 of 50,000 recombinant clones showed amylolytic activity. Subcloning and sequence analysis of a positive clone led to the identification a novel gene (amyPL) coding for α-am...

Soluble rat liver glucokinase was expressed at high levels at 22 degrees C in the BL21(DE3)pLysS strain of Escherichia coli. Aspartate-211 of yeast hexokinase has been implicated as a catalytic residue from crystallographic data. The corresponding residue in rat liver glucokinase, aspartate-205, was mutated to alanine and the expressed mutant had 1/500th of the activity of the wild type, with n...

UreG is one of the accessory proteins of urease in plants. In this study, a gene encoding an UreG consisting of 363 amino acids from Arabidopsis thaliana was isolated. The AtUreG gene was expressed in Escherichia coli BL21 as a GST fusion protein. The GST-AtUreG fusion protein was induced and purified under the optimized culturing conditions of 0.2 mM IPTG and 25°C for 5 h. Western blot analysi...

Salt-induced overexpression of genes cloned downstream of the phage T7 phi10 promoter was demonstrated in an Escherichia coli strain (GJ1158) which carries a single chromosomally integrated copy of the gene for phage T7 RNA polymerase under transcriptional control of the cis-regulatory elements of the osmoresponsive proU operon. Plasmids that have been constructed to obtain overproduction of in...

The canine monocytic ehrlichiosis, caused by Ehrlichia canis, is endemic in several regions of Brazil. Some serological diagnostic techniques using immunodominant proteins of E. canis as antigens are available, but their specificities and sensitivities are questionable. Based on this, the objective of this study was to test the antigenic potential of the recombinant gp19 protein (rGP19) for sub...