Optimization and One-Step Purification of Recombinant V Antigen Production from Yersinia pestis.

The purpose of this study was to develop an efficient and inexpensive method for the useful production of recombinant protein V antigen, an important virulence factor for Yersinia pestis. To this end, the synthetic gene encoding the V antigen was subcloned into the downstream of the intein (INT) and chitin-binding domain (CBD) from the pTXB1 vector using specific primers. In the following, the produced new plasmid, pTX-V, was transformed into E.

A new method for simultaneous gene deletion and down-regulation in Brucella melitensis Rev.1.

In this study, our aim was to integrate an antisense expression cassette in bacterial chromosome for providing a long-term expression down-regulation in a bid to develop a new approach for simultaneous deletion and down-regulation of target genes in bacterial system. Therefore, we were used this approach for simultaneous deletion of the perosamine synthetase (per) gene and down-regulation of the virB1 expression in Brucella melitensis Rev.1.

Stabilisation of recombinant aequorin by polyols: activity, thermostability and limited proteolysis.

The photoprotein aequorin is a calcium-dependent bioluminescent enzyme which is most widely used in biotechnology processes, but this protein is susceptible to aggregation and proteolysis degradation. Various additives such as polyols are known to enhance the stability of proteins and protect them in native folded and functional state. In this work, for study of aequorin stability, the histidine-tagged apoaequorin was expressed in Escherichia coli and purified by nickel chelate affinity chromatography.

Design and characterization of an aequorin-based bacterial biosensor for detection of toluene and related compounds.

An aequorin-based Escherichia coli strain JM109 biosensor was constructed and characterized for its potential to detect toluene and related compounds in aqueous solutions. The biosensor was constructed based on a PGL2 plasmid carrying the lower pathway promoter (Pu) of the xyl operon of Pseudomonas putida mt-2, which was incorporated with transcriptional activator xylR and fused to aequorin cDNA named pGL2-aequorin. Binding of xylR protein to a subset of toluene-like compounds activates transcription at the Pu promoter, thus expression of aequorin is controlled by xylR and Pu.