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Expression and characterization of recombinant human micro-plasminogen

Zhifeng Ma1, Wei Lu, Sheng Wu

  • 1Institute of Molecular Medicine, State Key Laboratory of Pharmaceutical Biotechnology, Nanjing University, Nanjing, China.

Biotechnology Letters
|January 9, 2007
PubMed

Insights

Researchers produced recombinant human micro-plasminogen (rh-microPlg) and found it activates faster with urokinase (UK) than Glu-plasminogen. The kringle 1-5 domain influences UK and plasmin activation processes.

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Protein Engineering

Background:

  • Plasminogen activation is crucial for fibrinolysis.
  • Understanding micro-plasminogen (microPlg) function requires efficient recombinant production.
  • The role of specific plasminogen domains in enzyme activation is not fully elucidated.

Purpose of the Study:

  • To express and purify recombinant human micro-plasminogen (rh-microPlg).
  • To compare the activation kinetics of rh-microPlg and Glu-plasminogen by urokinase (UK).
  • To investigate the catalytic efficiency of recombinant human micro-plasmin (rh-microPlm) and Lys-plasmin in pro-urokinase (proUK) activation.

Main Methods:

  • Reverse transcription PCR amplification of microPlg gene from human liver cells.
  • Insertion into pET-28a vector and transformation into E. coli BL21(DE3) for expression.
  • IPTG-induced over-expression, renaturation, purification, and enzymatic activity assays.

Main Results:

  • Successfully produced 16 mg/l of 95% pure rh-microPlg.
  • UK activation of rh-microPlg was significantly faster than Glu-plasminogen.
  • Lys-plasmin showed a fourfold greater catalytic efficiency in proUK activation compared to rh-microPlm.

Conclusions:

  • The kringle 1-5 domain of plasminogen/plasmin plays a role in modulating UK-mediated plasminogen activation.
  • This domain also influences plasmin-mediated proUK activation.
  • rh-microPlg is a valuable tool for studying plasminogen activation pathways.

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