BcLTP, a novel lipid transfer protein in Brassica chinensis, may secrete and combine extracellular CaM

Lipid transfer proteins in plants are believed to be involved in many processes of cell physiology and development. In this work, a full-length cDNA encoding a novel lipid transfer protein, designated BcLTP was isolated from Brassica chinensis. At least two copies of BcLTP are present in whole genome of B. chinensis, and its transcripts preferably accumulate in second-year organs, implying its role in reproductive growth stage. The 118 amino acid sequence deduced from a 354 bp open reading frame (ORF) shares common features with other members of plants LTPs family. A putative signal peptide at the N terminus was tested for secretion function by the yeast signal sequence trap (YSST) system, and further confirmed by vesicular and extracellular localization of YFP fusion protein. A highly conserved CaM binding site at C terminus was found and the binding properties with two representative CaM isoforms, one is convergent AtCaM2, one is divergent SCaM5, were determined by gel overlay. We found that convergent AtCaM2 prefer high concentration of Ca(2+) for binding BcLTP, while SCaM5 does not depend on Ca(2+ )concentration too much for binding BcLTP. The lipid binding feature of BcLTP was demonstrated using florescence-marked 1-pyrenedodecanoic acid, which can be enhanced by AtCaM2 in Ca(2+ )dependent manner and by SCaM5 in either presence or absence of Ca(2+). The collected data suggest that BcLTP may secrete and combine extracellular CaM isoforms, which in turn, facilitate lipid binding of BcLTP via Ca(2+) mediated signaling.