[Purification and functional identification of the recombinant human CREG/myc-His glycoprotein].

Zhongguo Ying Yong Sheng Li Xue Za Zhi

Department of Cardiology, Shenyang General Hospital, Cardiovascular Research Institute of PLA, Shenyang 110016, China.

Published: August 2010

AI Article Synopsis

  • The study aimed to purify and confirm the biological function of recombinant human cellular repressor of EIA stimulated gene (hCREG)/myc-His glycoprotein that inhibits the growth of smooth muscle cells from the human internal thoracic artery (HITASY) in vitro.
  • The hCREG/myc-His protein was purified using a Ni-NTA column and then desalted, achieving a concentration of 1.6 mg/ml with 92% purity; it was also identified as glycosylated.
  • Results showed that lower concentrations of the hCREG protein (0.5 and 1 microg/ml) were more effective in inhibiting HITASY cell proliferation compared to higher concentrations (2 microg

Article Abstract

Objective: To purify the recombinant human cellular repressor of EIA stimulated gene (hCREG)/myc-His glycoprotein and confirm the biological function of hCREG/myc-His which could inhibit the proliferation of human internal thoracic artery smooth muscle cells (HITASY) cultured in vitro.

Methods: The recombinant hCREG/myc-His protein was purified with Ni-NTA column according to 6 x His affinity chromatographic theory. The recombinant hCREG/myc-His protein was desalted by HiTrap Desalting Column. The effect of recombinant hCREG/myc-His glycoprotein of different concentration (0.5 microg/ml, 1 microg/ml and 2 microg/ml) on proliferation of HITASY cells was studied by flow cytometric analysis and the effect of recombinant protein on proliferation of HITASY cells was confirmed by BrdU incorporation method.

Results: The recombinant hCREG protein was purified with Ni-NTA column according to 6 x His affinity chromatographic theory. The concentration of recombinant hCREG protein which has been concentrated and desalted was determined to be 1.6 mg/ml and the purity of recombinant protein reached 92%. The protein was identified to be glycosylated. The recombinant hCREG protein was identified to inhibit the proliferation of HITASY cells cultured in vitro and the inhibition effect was stronger in low-dosage group than that in high-dosage group by flow cytometric analysis. The proliferation of HITASY cells cultured in vitro with 2 microg/ml recombinant hCREG protein was inhibited significantly compared with that in control group according to the BrdU incorporation result. There was statistical difference among the groups (P < 0.05).

Conclusion: The purification of recombinant hCREG/myc-His glycoprotein with biological activity provides an experiment platform for function study and engineering production of hCREG protein.

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