Biochemical Features of Recombinant Human Cyclophilin J

Anticancer Res. 2016 Mar;36(3):1175-80.

Abstract

Aim: To characterize the biochemical features of the newest member of cyclophilin family of peptidyl-prolyl cis/trans-isomerases (PPIases), cyclophilin J (CYPJ).

Materials and methods: PPIase assays were performed on purified hCYPJ and its mutated variants. The substrate specificity, half-maximal inhibitory concentration (IC50) of cyclosporin A (CsA) inhibition and circular dichroism (CD) spectrum of CYPJ were measured. Mercury pathway profiling luciferase assays were also performed.

Results: The catalytic number/Michaelis constant (kcat/KM) value of CYPJ was 9.5×10(4) s(-1)M(-1). CYPJ additionally catalyzed norleucine-proline, isoleucine-proline and glutamine-proline peptides compared to CYPA and Escherichia coli PPIases. CYPJ was inhibited by CsA in a dose-dependent manner with IC50 of 12.1±0.9 μM. The CD spectrum of CYPJ was similar to CYPA. CYPJ significantly up-regulated the transcription of E-box, E2F, retinoblastoma (Rb), p53, activator protein 1 (AP1), NF-κB and phospho-cAMP response element (CRE) cis-response element in 293T cells.

Conclusion: CYPJ structurally resembles CYPA. It is sensitive to inhibition by CsA and plays a role in regulating cell growth, proliferation, and apoptosis.

Keywords: CYPJ; Cyclophilin J; PPIL3; PPIase activity; biochemistry.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Catalysis
  • Cyclic AMP Response Element-Binding Protein / genetics
  • Cyclic AMP Response Element-Binding Protein / metabolism
  • Cyclophilins / antagonists & inhibitors
  • Cyclophilins / genetics
  • Cyclophilins / metabolism*
  • Cyclosporine / metabolism
  • Cyclosporine / pharmacology
  • Dose-Response Relationship, Drug
  • E2F Transcription Factors / genetics
  • E2F Transcription Factors / metabolism
  • Enzyme Inhibitors / metabolism
  • Enzyme Inhibitors / pharmacology
  • Gene Expression Regulation
  • HEK293 Cells
  • Humans
  • Kinetics
  • Mutation
  • NF-kappa B / genetics
  • NF-kappa B / metabolism
  • Protein Binding
  • Recombinant Proteins / metabolism
  • Retinoblastoma Protein / genetics
  • Retinoblastoma Protein / metabolism
  • Substrate Specificity
  • Transcription Factor AP-1 / genetics
  • Transcription Factor AP-1 / metabolism
  • Transcription, Genetic
  • Transfection
  • Tumor Suppressor Protein p53 / genetics
  • Tumor Suppressor Protein p53 / metabolism

Substances

  • CREB1 protein, human
  • Cyclic AMP Response Element-Binding Protein
  • E2F Transcription Factors
  • Enzyme Inhibitors
  • NF-kappa B
  • Recombinant Proteins
  • Retinoblastoma Protein
  • TP53 protein, human
  • Transcription Factor AP-1
  • Tumor Suppressor Protein p53
  • Cyclosporine
  • Cyclophilins
  • PPIL3 protein, human