[Molecular cloning, expression, purification and properties of isocitrate lyase gene from Mycobacterium tuberculosis H37Rv].

Objective To obtain recombinant protein with enzymatic activities of isocitrate lyase(ICL).Methods The icl gene was amplified by polymerase chain reaction(PCR) from Mycobacterium tuberculosis H_(37)Rv strain genomic DNA and cloned into pET28-a(+) vector.The recombinant protein was expressed in E.coli BL21(DE3).Enzyme activity of the protein was assayed after purifing with Ni-NTA resin.Results The recombinant ICL was purified in a highly active state with a specific activity of about 7.657×10~2 μmol·mg~(-1)·min~(-1).The pH curve indicated that recombinant ICL activity was optimal at pH 7.4.The LC/MS spectrometry showed a 50 603.347 molecular mass of recombinant ICL.The CD spectrum showed that the percentages for α-helix,β-sheet, β-turn,and random coil were 43.8 %,31.9%,3.4%,and 20.9 %,respectively.Conclusions The icl gene of Mycobacterium tuberculosis H_(37)Rv was successfully cloned and expressed.The enzymatic properties demonstrated the purified recombinant protein had activities of ICL.This work can facilitate immunologic research and the discovery of novel antimicrobial agents against Mycobacterium tuberculosis.