The low abundance and highly hydrophobic nature of most membrane proteins make their analysis more difficult than that for common soluble proteins.Successful membrane protein identification is largely dependent on the sample preparation including the enrichment and dissolution of the membrane proteins.A series of conventional and newly developed methods has been applied to the enrichment of low-abundance membrane proteins at membrane and/or protein levels and to the dissolution of hydrophobic membrane proteins.However,all the existing methods have inherent advantages and limitations.Up to now,there has been no unique method that can universally be employed to solve all the problems and more efforts are needed in improving sample preparation for the analysis of membrane proteomes.
Human trefoil factor 2 （hTFF2） is considered as one of the most important initiators of mucosal healing in the gastrointestinal tract by promoting cell migration and suppressing apoptosis. However, it is hard to obtain hTFF2 from human tissue and many recombinant hTFF2 produced in vitro exist as fusion proteins. The purpose of the present study was to produce native hTFF2 while maintaining its biological activities. The open reading frame of hTFF2 was inserted into a pET-32a（＋） expression vector, and hTFF2-TRX fusion protein was successfully expressed in Escherichia coli and purified by Nickel-nitrilotriacetic acid affinity chromatography and reverse-phase HPLC steps. The recombinant fusion protein （purity〉95%） was cleaved by Factor Xa at 23 ~C to release hTFF2. After removal of Factor Xa and undigested fusion proteins, hTFF2 was purified and identified by SDS-PAGE and Western blotting. The yield of recombinant hTFF2 was about 5 mg/L. The recombinant hTFF2 could promote IEC-6 cells migration and in vitro wound healing via the activation of ERK1/2. Recombinant hTFF2 could also inhibit apoptosis of HCT-116 cells induced by 50 lamol/L ceramide In summary, our results showed that the recombinant hTFF2 was expressed in E. coli and successfully purified after cleavage with the fusion partner with high yield while maintaining its biological activities. Recombinant hTFF2 might be useful for investigating the molecular mechanism of hTFF2 and development of hTFF2-related drugs.
This review describes the history of taxonomic research on scorpions and provides an updated checklist and key of the scorpions currently known in China. This checklist is based on a thorough review of the extant literatures on scorpion species whose presence has been confirmed in China through field expeditions and examination of scorpion collections, excepting a few members that have no clear distribution or are currently in doubt. Totally, the scorpion fauna of China consists of 53 species and subspecies belonging to 12 genera crossing five families, with 33 species(62.3%) and one genus being recorded as endemic. Additionally, identification key and the distribution of scorpions from China are provided.
Zhi-Yong DIZi-Zhong YANGShi-Jin YINZhi-Jian CAOWen-Xin LI
The present study was designed to identify immunomodulatory components from the leech salivary gland of Haemadipsa sylvestris. The Sephadex G-50, ResourceTM S column chromatography and reverse-phase high performance liquid chromatography(RP-HPLC) were used to isolate and purify the salivary gland extracts(SGE). Structural analysis of isolated compounds was based on Edman degradation and matrix assisted laser desorption ionization time-of-flight mass spectrometer(MALDI-TOF-MS). The c DNA encoding the precursor of the compound was cloned from the c DNA library of the salivary gland of H. sylvestris. The levels of inflammatory mediators, including tumor necrosis factor-α(TNF-α), interferon ?(IFN-?), interleukin-6(IL-6), and monocyte chemotactic protein-1(MCP-1) were assayed using an enzyme-linked immunosorbent assay(ELISA). The effects on cell proliferation and cell viability were observed using MTT assay. A novel neuropeptide Y(Neuropeptide Y-HS) from the leech salivary gland of H. sylvestris was purified and characterized. It was composed of 36 amino acid residues and the amino acid sequence was determined to be FLEPPERPAVFTSVEQMKSYIKALNDYYLLLGRPRF-NH2, containing an amidated C-terminus. It showed significant inhibitory effects on the production of inflammatory cytokines including TNF-?, IFN-?, IL-6, and MCP-1. Neuropeptide Y was identified from leeches for the first time. The presence of neuropeptide Y-HS in leech salivary gland may help get blood meal from hosts and inhibit inflammation.
LIU Wei-HuiCHEN YanBAI Xue-WeiYAO Hui-MinZHANG Xu-GuangYAN Xiu-WenLAI Ren
