[Purification and gene cloning of a novel fibrinolytic protease from Streptomyces sp. C3662].
A novel fibrinolytic protease from Streptomyces sp. C3662 was purified by (NH4)2SO4 fractionation, DEAE-Sepharose and CM-Sepharose chromatography. The molecular weight of the protease was indicated to be 30 kD by SDS-PAGE. Using a E. coli/Streptomyces shuttle plasmid pIJ699 as the vector, shot-gun cloning was performed to clone the gene of the protease. One clone with fibrinolytic activity harboring a plasmid that contains a DNA fragment of 6 kb was obtained from 3000 clones. Sequence analysis reveals that the open reading frame of the gene of the protease is 903 bp in size, encoding a putative protein of 300 amino acids with 30 kD. The overall GC% and the third codon GC% of the ORF were 68.33% and 95.6%, respectively. Comparison of homologue showed that the purative protein is highly homologous with other proteases of Streptomyces sp.
Gong
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Y
Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences, Peking Union Medical College, Beijing 100050, China.
Wang
Y
Y
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics*; D010450:Endopeptidases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016366:Open Reading Frames / Q000235:genetics*; D017385:Sequence Homology; D013302:Streptomyces / Q000201:enzymology* / Q000235:genetics
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D010450:Endopeptidases