96% and with the same bio-activity as unlabeled Huwen toxin-1; Radioactivity detected in epidural space was 38% of injected radioacti vity at 10 min after epidural injection, which demonstrated successful administr ation into epidural space; The maximum serum concentration after epidural or iv administration of [ 125 I]labeled Huwentoxin-1 were determined to be (0 70?0 04) MBq?L -1 and (4 98?0 58) MBq?L -1 , respectively, a t the maximum serum concentration times of 30 min and 2 min. Terminal T 1/2 after epidural or iv administration were (10 36?0 27) h or (11 03?1 16) h, respectively. Cls was (1 29?0 07) L?h -1 ?kg -1 or (1 25? 0 23) L?h -1 ?kg -1 , respectively. Bioavailability after epidural a dministration was(95?5)%. CONCLUSION Concentration-time cur ves of [ 125 I] labeled Huwentoxin-1 after two routes were different. The degradation profiles after epidural and iv injection supported the using of HWTX-1 as analgesic by epidural administration.

Rv2742 is a novel gene identified from Mycobacterium tuberculosis H37Rv by the proteogenomics strategy. The aim of this study was to establish a system of soluble expression and purification of the missing protein Rv2742 in M. tuberculosis H37Rv, to provide reference for further research on the biological function of Rv2742. The soluble protein was not successfully induced by prokaryotic expression vectors pGEX-4T-2-Rv2742, pET-32a-Rv2742, pET-28a-Rv2742 and pMAL-c2X-Rv2742. After the codon of novel gene Rv2742 was optimized according to E. coli codon usage frequency, only the recombinant strain containing plasmid pMAL-c2X-Rv2742 could produce soluble products of Rv2742 encoding gene. In addition, the expression effects of the desired fusion protein were also analyzed under different conditions including hosts, culture temperatures and IPTG concentrations. The optimum expression conditions were as follows: Rosetta (DE3) host, 16 °C culture temperature and 0.5 mmol/L IPTG. After being purified by affinity chromatography with amylose resin, the fusion protein sequence was confirmed by LC-MS/MS. These results indicated that the novel gene Rv2742 product could be successfully induced and expressed in a soluble form by the expression system pMAL-c2X with MBP tag. Our findings provide reference for studies on potential interaction and immunogenicity.
Chromatography, Liquid ; Cloning, Molecular ; Escherichia coli ; Mycobacterium tuberculosis ; genetics ; Recombinant Fusion Proteins ; Tandem Mass Spectrometry