A multi-functional zinc(II) complex of an imidazolium ionic liquid-tagged ligand was synthesized and characterized. The DSC and TGA analysis showed the thermal stability of the complex. The complex showed emission λmax at 368–467 nm with quantum yields up to 0.06. The time-resolved fluorescenc...

Exotoxin A (ETA) has been described as a major virulence factor produced by the opportunistic pathogen Pseudomonas aeruginosa. The transcription of the ETA structural gene (toxA) has been shown to be positively regulated by the product of the toxR gene (also called regA). However, the mechanism ...

ToxR is a transmembrane, DNA-binding protein that can activate transcription of genes encoding cholera toxin (ctxAB). Here we characterize ToxS, a 19 kd transmembrane regulatory protein that interacts with ToxR and stimulates its activity. If a portion of the periplasmic domain of ToxR is delete...

PCR protocols directly applied to enrichment broth cultures were compared with a culture method based on the ISO reference for detection of Vibrio parahaemolyticus in 57 natural bivalve mollusc samples. Comparisons were made on different primer pairs specifically targeting the V. parahaemolyticu...

The toxR gene encodes a transcriptional activator controlling cholera toxin, pilus, and outer-membrane protein expression in V. cholerae. Nucleotide sequence and mutational analysis has identified the toxR gene product as a 32,527 dalton protein. Hydropathicity analysis of the derived amino acid...

Specific interactions between membrane spanning polypeptide segments are important for folding and oligomerisation of integral membrane proteins. Previously the dimerisation of glycophorin A has been shown to depend on interactions between its transmembrane segment by studying chimeric proteins ...

The transcription activator ToxR controls the expression of cholera toxin, pilus colonization factor and outer membrane protein in Vibrio cholerae. It binds to the 5′-TTTTGAT-3′ tandemly repeated DNA sequence in the cholera toxin promoter region. ToxR is a membrane protein having distinct peri...

Assay systems based on the ToxR protein are widely used to investigate interaction of transmembrane domains that come from natural proteins or are isolated from combinatorial libraries. The principle of this method is that self-interaction of any given transmembrane domain, which is expressed wi...

ToxR-based transcriptional reporter assays allow the strength of transmembrane helix interactions in biological membranes to be measured. Previously, these assays have only been used to study single-pass transmembrane systems. To facilitate investigation of polytopic transmembrane domain (TMD) o...

In order to assess the potentiality of Vibrio cholerae ToxR protein and of bacteriophage λ repressor as indicators of the dimerization of periplasmic proteins in Escherichia coli, we have constructed a series of plasmids encoding transmembrane fusion proteins. The amino-terminal part, containin...

Vibrio parahaemolyticus is a seafood-borne Gram-negative bacteria causing diarrheal diseases in humans world wide. ToxR is a membrane-associated transcriptional factor which plays an important role in acid stress tolerance and regulates the expression of virulence genes including type III secret...

Hirudin (HIR), derived from leeches, and tick anticoagulant peptide (TAP) are polypeptide protease inhibitors of thrombin and coagulation factor Xa (fXa), respectively, and they have both shown utility in vitro and in vivo as potent antithrombotic agents. A thorough side-by-side comparison of th...

The scene of the protein micro-heterogeneity of recombinant hirudin-II (HV2) expressed in Pichia pastoris was investigated. It was shown that three derivatives of HV2 were present in the fermentation broth of P. pastoris, which were intact HV2 and its two derivatives truncated the C-terminal ami...

Serious loss of polyethylene glycol (PEG) reagent is confronted during PEGylation of proteins in mildly acidic aqueous solution. In this study, a novel approach of PEGylation of recombinant hirudin variant-2 (HV2) with monomethoxy-PEG-succinimidyl carbonate (mPEG-SC) was developed in six mixed a...

Recombinant Neorudin (EPR-hirudin, EH), a novel, low-bleeding anticoagulant fusion protein, has been developed as an inactive prodrug that is converted to an active metabolite, hirudin variant 2-Lys47 (HV2), at the thrombus site and is undergoing Phase I clinical trials in China. The goal of our...

In this study, an integrated process was developed for successive solid-phase PEGylation of recombinant hirudin variant-2 (HV2) and separation of PEGylated HV2 species on an anion exchange chromatography column (so-called in situ PEGylation). The effects of different PEG sizes, ion exchange resi...

To improve the therapy efficacy of recombinant hirudin variant-2 (HV2), its PEGylation was investigated using linear mPEG-succinimidyl carbonate (mPEG-SC) and branched mPEG2-N-hydroxysuccinimide (mPEG2-NHS). The reaction mixtures of PEGylation were analyzed by RP-HPLC and the mono-PEG-HV2 produc...

A critical challenge of PEGylation is the production of the desired PEGylated protein form at a high yield. In this study, a kinetic model was constructed successfully to describe the PEGylation reaction of recombinant hirudin variant-2 (HV2) with monomethoxy-PEG-succinimidyl carbonate (mPEG-SC)...

In this study, chemical modification of recombinant hirudin variant-2 (HV2) with palmitic acid (PAL) was proposed as an alternative approach to circumvent the limitations of PEGylation. To facilitate a sufficient contact of the hydrophilic HV2 to the hydrophobic PAL, thereby improving the reacti...

The melanocortin subtype-4 receptor (MC4R) has been implicated in the control of feeding behavior and body weight regulation. A series of tetrapeptides, based on Tic-DPhe-Arg-Trp-NH2—a mimic of the putative message sequence “His-Phe-Arg-Trp” and modified at the DPhe position, were prepared an...