Characterization of a novel Cyclophilin-type peptidylprolyl isomerase protein from sweet potato storage roots

An antioxidant protein of cyclophilin-type peptidylprolyl isomerase (SPPPI) from sweet potato (Ipomoea batatas (L.) Lam. 'Tainong 57') storage roots was isolated by differential display. The open reading frame of this cDNA encodes a pro-protein of 260 amino acids with a predicted molecular mass of 27,658 Da (pI 9.34). A comparison of the deduced amino acid sequence of SPPPI with precursor proteins indicates 65% identity to the Arabidopsis thaliana AraPPI sequence. Computer analysis of the deduced amino acid sequences of the conserved domain revealed that the protein belonged to the plant cyclophilin-type peptidylprolyl isomerase. Genomic Southern blot analyses using the full-length SPPPI cDNA probe revealed a multigene family in the sweet potato genome. Both the corresponding mRNA and protein level were found the highest in the storage roots, followed by that in sprout. Recombinant SPPPI overproduced in E. coli (M15) was purified by Ni²⁺-chelated affinity chromatography. Both the peptidylprolyl isomerase and antioxidantive activity of active recombinant SPPPI were investigated. SPPPI and CP (calf thymus cytophilin, a positive control) displayed the highest ABTS (2, 2-azino-bis-(3-ethylbenzothiazoline- 6-sulfonic acid) scavenging ability (15.36 ± 0.80 and 17.79 ± 1.72%, respectively) at 100 μg/mL. In the DPPH (1, 1-diphenyl-2- picrylhydrazyl) assay, SPPPI and CP were found to have the highest radical- scavenging activity (5.78 ± 0.62 and 4.05± 0.80%, respectively) at 100 μg/mL. The Fe²⁺-chelating ability of SPPPI and CP was found to be the highest (12.47± 2.37 and 14.57± 0.96%) at 100 μg/mL, respectively. It was suggested that SPPPI is an excellent candidate as a lead compound for the development of reductant agents.