Project's information

Project's title Cloning, high-level expression and orientated application of the recombinant streptokinase a clot-buster drug in medicine
Research hosting institution Institute of Biotechnology
Project leader’s name Prof. Dr. Quyen Dinh Thi
Project duration 01/01/2008 - 01/01/2009
Project’s budget 300.000.000 VND
Classify Excellent
Goal and objectives of the project

Creat product of streptokinase to treat thrombosis by drinking way.
Main results

Scientific results

  • Four Streptococcus strains including S. pyogenes, S. equisimilis, S. uberis, Staphylococcus aureus were isolated and two genes encoding streptokinase from S. pyogenes and S. equisimilis. was cloned.
  • SK gene from S. pyogenes was inserted into the expression vector pET22b+ and expressed in E. coli BL21 (SKBL21) at a level of 7591 U/ml culture medium with a specific activity of 11264 U/mg protein.
  • SK gene from S. pyogenes was inserted into the expression vector pPICZαA and expressed in P. pastoris at a level of 2374 U/ml culture medium.
  • Recombinant SK was purified through affinity chromatography column of Ni2+ - ProBondTM resin and investigated on some physicochemical properties.
  • Production yield of purified SK from SKBL21 was 2 mg protein/g cell.
  • Recombinant SK showed no toxicity on mice and thrombolysis on rabbit model (187500 to 225000 U/mouse).
Novelty and actuality and scientific meaningfulness of the results

SK production in Vietnam was new.
Products of the project

Scientific papers in referred journals (list):

  • Vu Hong Diep, Nguyen Thi Ngoc Dao, Quyen Dinh Thi (2008): Cloning and sequence analysis of a gene encoding streptokinase from Streptococcus pyogenes DT7. Journal of Biotechnology 6(4A): 757-763.
  • Nguyen Thi Thao, Vu Hong Diep, Quyen Dinh Thi et al (2009): Generation of genetically modified microoganisms producing recombinant enzyme for agricultural, industrial and midical applications. Proceedings of the Natl Conf Genet Modif Microorg Manag Biosaf, Hanoi 8/2009: 115-126.
  • Vu Hong Diep, Quyen Dinh Thi et al. (2009): Identification of some hemolytic bacteria isolated in Vietnam by 16S rRNA gene analysis. Proceedings of Natl Conf Biotechnol, Thai Nguyen 11/2009: 65-68.

Patents (list): “Recombinant Escherichia coli strain BL21 producing streptokinase”, register number 1-2010-00803.

  • Technological products (describe in details: technical characteristics, place):
  • 03 recombinant E. coli strains BL21 producing high-level of streptokinase with expression vector pET22b+, stored at -80°C, Dept of Enzyme Biotechnology.
  • 01 recombinant P. pastoris strain producing high-level of SK with expression vector pPICZαA, stored -80°C, Dept of Enzyme Biotechnology.
  • 100 mg purified SK power, stored at 4°C, Dept of Enzyme Biotechnology.
Recommendations

Further purification of SK to use as thrombosis agent by injection way.