Heterologous expression,chaperone mediated solubilization and purification of parasitic nematode-specific growth factor-like protein of Setaria digitata

作     者：W.WP.Rodrigo R.S Dassanayake E.H.Karunanayake Y.I.N.Silva Gunawardene O.VDS.J.Weerasena 

作者机构：Department of ChemistryFaculty of ScienceUniversity of Colombo90Cumaratunga Munidasa Mawatha.Colombo 03Sri Lanka Institute of BiochemistryMolecular Biology and BiotechnologyUniversity of Colombo90Cumaratunga Munidasa MawathaColombo 03Sri Lanka Molecular Medicine UnitFaculty of MedicineUniversity of Kelaniya 

出 版 物：《Asian Pacific Journal of Tropical Medicine》 (亚太热带医药杂志（英文版）)

年 卷 期：2014年第7卷第2期

页      面：85-92页

核心收录：

学科分类：090601[农学-基础兽医学] 1002[医学-临床医学] 09[农学] 0906[农学-兽医学] 

基　　金：supported by a grant(SIDA/2006/BT/04) awarded by National Science Foundation of Sri Lanka 

主　　题：Setaria digitata Uncharacterized gene Chaperon Co-expression 

摘      要：Objective:To clone,express and purify a putative parasitic nematode specific protein of Setaria digitata(***),filarial nematode that infects livestock and cause significant economic losses in Far East and Asia to he used for structural and functional ***:To characterize uneharacterized gene of,***(SDUG),the herterologous expression of SDUG was carried out in the pET[cloned into pET45b(+)]expression system initially and co-expression of SDUC using chaperoiie plasmids pG-KJE8,pGro 7,pKJE7,pG-Tf2 and pTf16 containing chapcrone proteins of dnaK-dnaJ-grpE-groES-gro-E,groES-groEL,dnaK-dnaJ-grpE,groES-groEL-tig,and tig respectively,was carried out ***:Expression of SDUG was seen when Escherichia coli strain BI.21(DE3)is used,while concentrating protein largely into the insoluble *** co-expression of SDUG using chaperoiie plasmid mediated system indicated a significant increase of the protein in the soluble *** the chaperon plasniid sets,the highest amount of recombinant SDUP in the soluble fraction was seen when pGro7 was used in the presence of2 mg/mL L-arabinosc and 0.6M IPTG concentration in the culture medium and for 3 h of incubation at the temperature of 28℃.Recombinant SDUG was purified both from soluble and insoluble fractions using Ni affinity ***-PAGE and western blot analyses of these proteins revealed a single band having expected size of^24 ***:SDUG seems to be more aggregate-prone and hydrophobic in nature and such protein can make soluble by correct selecting the inducer concentrations and induction temperature and its duration.