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