Characterization of bifunctional chimeric molecule of PRGDWR containing pro-urokinase

Xin Dang, Jing Xin Yang, Qiang Ru, Bing Gen Ru*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

Abstract

In order to obtain the bifunctional chimeric molecule of single-chain urokinase-type plasminogen activator (scu-PA) which can inhibit platelet aggregation, PRGDWR peptide was inserted into the site between Gly118 and Leu119 (called insertion mutant B, InB). The recombinant gene of InB was expressed by Pichia pastoris. The secreted protein was purified by metal chelate affinity and strong cation exchange chromatography. The amidolytic ability of mutant InB is 5 900 IU/mg, the kinetic constants is: Km,plgInB = 56.8 μmol·L-1, kcat,plgInB = 0.33 s-1. The kinetic constants of plasminogen activation reaction is: Km,plgInB = 0.397 μmol·L-1, kcat,plgInB = 0.0164 s-1. Fibrin inhibit the catalytiv ability of InB during plasminogen activation, the influence factor is 0.463 (means InB remain 46.3% of the catalytic ability when fibrin was involved in the reaction system). The mutant not only has almost the same catalytic ability as wild type scu-PA, but also has strong ability of anti-platelet aggregation (compared with scu-PA), IC50 of InB is 12.7 μmol·L-1.

Original languageEnglish (US)
Pages (from-to)208-209
Number of pages2
JournalProgress in Biochemistry and Biophysics
Volume28
Issue number2
StatePublished - Dec 1 2001
Externally publishedYes

Keywords

  • PRGDWR peptide
  • Platelet aggregation
  • Single chain urokinase-type plasminogen activator
  • Thrombolytic ability

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry

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