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CLONING  AND  PROKARYOTIC  EXPRESSION  OF  A COMPLEMENTARY DNA GENE FOR CYCLOPHILIN FROM CAMELLIA OLEIFERA

XIAOFENG TAN^1*, BAOMING WANG¹, LIN ZHANG¹, MINGWANG SHI^{1, 2}, LUSHAN XIE¹ AND FANGMING HU¹

Abstract: Cyclophilins (CyPs) are ubiquitous proteins and involved in protein folding and stress response due to peptidyl-prolyl cis-trans isomerase (PPIase) activity. In this study, a 975-base pair (bp) full-length complementary DNA (cDNA) gene, Camellia oleifera CyP (CoCyP, GenBank access no. FJ377540), was isolated from the conducted cDNA library of C. oleifera seeds. This cDNA encodes a polypeptide of 207 amino acids, which has an endoplasmic- reticulum-localized signal sequence at its N-terminus, but not the typical signal sequence at its C-terminus. Moreover, its putative amino acid sequence shares highest identity with those of Arabidopsis thaliana CyPB (84%) and Triticum aestivum CyPB (81%), and possesses a highly conserved central core domain including cyclosporin A (CsA)-binding sites, the tetrapeptide Ala-Ala-Pro-Ala-binding sites, residues required for PPIase catalysis and an insertion of seven amino acids of unknown function. Collectively, it is suggested that the CoCyP protein belongs to CyPB and has PPIase activity.  An approximately 21 kDa protein was expressed via the recombinant pET-30b(+)/CoCyP in Escherichia coli. In this study, we tentatively put forward to the hypothesis that the gene expressing during the peak of seed lipid biosynthesis, might correlate with the caused harmful products in the lipid biosynthesis process, and protect cells against reactive oxygen species (ROS) damage; thus it may be crucial during lipid biosynthesis and stress responsiveness.

¹The Key Lab. of Non-wood Forest Products of State Forestry Administration,

Central South University of Forestry and Technology, Changsha, 410004, P. R. China

²Henan Institute of Science and Technology, Xinxiang, 453003, P. R. China

^*Corresponding author Email: tanxiaofengcn@126.com; Tel: +86 731-5623416