Production, purification of the recombinant analog of y-box-binding protein 1 and its interaction with poly(ADP-ribose), RNA, single- and double-stranded DNAs

E. E. Alemasova, K. N. Naumenko, P. E. Pestryakov, O. I. Lavrik

Research output: Contribution to journalArticlepeer-review

2 Citations (Scopus)

Abstract

Aim. Production and purification of the recombinant histidine-tagged Y-box- binding protein 1 and study of its interaction with DNA, RNA and poly(ADP-ribose). Methods. Ligationindependent cloning, PCR, Sanger sequencing, protein chromatography, polyacrylamide gel electrophoresis, and electrophoresis mobility shift assay. Results. cDNA coding for the YB-1 protein has a previously undocumented two single nucleotide polymorphisms. The expression construct for production of the his-tagged YB-1 protein was designed to simplify the purification procedure and an appropriate protocol for protein purification was developed. Using electrophoresis mobility shift assay, we have shown that poly(ADP-ribose) competes with a double- and single-stranded DNA and RNA for binding to purified recombinant his-tagged YB-1. Conclusions. In the present work we developed and optimized the procedure of the recombinant YB-1 protein production and purification from bacterial cells. We found that poly(ADP-ribose) at high concentration is able to recruit YB-1 protein from the YB-1-DNA and YB-1-RNA complexes, suggesting a possible YB-1 involvement in DNA repair.

Original languageEnglish
Pages (from-to)214-220
Number of pages7
JournalBiopolymers and Cell
Volume33
Issue number3
DOIs
Publication statusPublished - 2017

Keywords

  • DNA repair
  • Poly(ADP-ribose) (PAR)
  • Protein purification
  • YB-1

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