NDLI: Pitfalls in screening streptococci for retrieving superior streptokinase (SK) genes: no activity correlation for streptococcal culture supernatant and recombinant SK

Content Provider	Springer Nature Link
Author	Keramati, Malihe Roohvand, Farzin Aslani, Mohammad Mehdi Motevalli, Fatemeh khatami, Shohreh Memarnejadian, Arash
Copyright Year	2012
Abstract	Streptokinase (SK), the heterogeneous protein family secreted by some groups of β-hemolytic streptococci (βHS), is a plasminogen activator and well-known drug for thrombolytic therapy. Differences in plasminogen activation property of streptococcal culture supernatants (SCS) have been traditionally used to identify superior producer strains and SK genes (skc) for recombinant SK (rSK) production. However, the role of SK heterogeneity and whether SK activities in SCS correlate with that of their corresponding rSK is a matter of debate. To address these concerns, SCS of nine group C streptococci (GCS) screened among 252 βHS clinical isolates were compared for plasminogen activation using S-2251 chromogenic assay. The GCS (Streptococcus equisimilis) showing the highest (GCS-S87) and lowest (GCS-S131) activities were selected for PCR-based isolation of skc, cloning and rSK production in Escherichia coli. The 6×His-tagged rSK proteins were purified by NI–NTA chromatography, analyzed by SDS-PAGE and Western blotting and their activities were determined. While SCS of GCS-S87 and GCS-S131 showed different plasminogen activations (95 and 35 %, respectively) compared to that of the reference strain (GCS-9542), but interestingly rSK of all three strains showed close specific activities (1.33, 1.70, and 1.55 × 104 IU mg−1). Accordingly, SKS87 and SKS131 had more than 90 % sequence identity at the amino acids level compared to SK9542. Therefore, SK heterogeneity by itself may not contribute to the differences in plasminogen activation properties of SCS and evaluation of this activity in SCS might not be a proper assay for screening superior skc.
Starting Page	151
Ending Page	158
Page Count	8
File Format	PDF
ISSN	13675435
Journal	Journal of Industrial Microbiology and Biotechnology
Volume Number	40
Issue Number	1
e-ISSN	14765535
Language	English
Publisher	Springer-Verlag
Publisher Date	2012-10-25
Publisher Place	Berlin, Heidelberg
Access Restriction	Subscribed
Subject Keyword	Plasminogen activation Plasmin amidolytic substrate Recombinant streptokinase Streptococcus equisimilis Biochemistry Inorganic Chemistry Genetic Engineering Biotechnology Bioinformatics Microbiology
Content Type	Text
Resource Type	Article
Subject	Medicine Bioengineering Applied Microbiology and Biotechnology Biotechnology

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