Expression,
Purification and Function of Rice Nonspecific Lipid Transfer Protein
GE Xiao-Chun*, CHEN Ji-Chao,
LIN Yi, SUN Chong-Rong, CAO Kai-Ming
( Department of Biochemistry, College of Life Sciences, Fudan University,
Shanghai 200433, China )
Abstract Plant
nonspecific lipid transfer protein(nsLTP) is a class of protein which has in
vitro lipid transferring activity between biomembranes. In order to study
the antimicrobial function of rice nonspecific lipid transfer protein, a gene LTP110
encoding rice nsLTP was cloned into ThioFusionTM expressionvector
pET32a(+) and expressed in host strain Bl21(DE3)trxB–. After induction
by IPTG at 30 °C for 5 h, the fusion protein thio-LTP110 was in large amount
produced. The expressed protein was purified by Ni2+-chelating
Sepharose fast flow column, then digested by enterokinase. By passing through
nickel affinity column again, the cleavage product, LTP110, was obtained. CD
spectrum scanning from 185 nm to 250 nm showed that the recombinant protein
LTP110 had similar secondary structure with the nsLTP purified from rice
etiolated seedlings. Activity determination by fluorescent lipid P-96 showed
that it had lipid binding activity. Microbial inhibition test results revealed
that LTP110 deterred germination of the spores of rice pathogen P.oryzae,
showing it might be involved in plant microbial resistance function. Therefore,
it has the potential to be used in plant transgene engineering to improve plant
resistance.
Key words nonspecific lipid transfer protein; lipid
binding activity; resistance function; P.oryzae
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