Molecular cloning and characterization of a cytoplasmic cyclophilin gene in sugarcane

Abstract

Cyclophilins are ubiquitous proteins with an enzymatic activity of peptidyl-prolyl cis-trans isomerase (PPIase), which play important roles in a variety of stress responsiveness. In this study, we reported the cloning and characterization of a full-length cytoplasmic cyclophilin gene in sugarcane. Sequence analysis showed the cDNA of this gene (GenBank accession number:GQ246462), termed as Sc-CyP, was 904 bp long, including a 519 bp complete ORF, the 5’ UTR of 74 bp and 3’UTR of 311 bp, plus a typical AATAA motif and poly (A) tail. It encoded the 172 amino acid polypeptide with a molecular weight of 18.4 KD and the isoelectric point of 8.68. The Sc-CyP encoding protein had the conserved site Trp128 (W128) ubiquitious of all cyclophilins in eukaryotes and the KSGKPLH48-54 region specific to cytoplasmic cyclophilins in plants. SDS-PAGE analysis and PPIase assay revealed that the expression product, with PPIase activity, was a fusion protein with a molecular weight about 25 and 18.4 kD of Sc- CyP plus 7 kD of His • Tag peptides. In real-time qPCR analysis, the Sc-CyP gene showed induced expression under PEG, NaCl, SA and H2O2 stresses, indicating it a stress-related gene for drought and salt stress, signal transduction and disease resistance response in sugarcane.

Key words: Sugarcane (Saccharum officinarum), cyclophilin, PPIase, real-time quantitative PCR.