Optimization of high expression and purification of recombinant streptokinase and in vitro evaluation of its thrombolytic activity was written by El-Dabaa, Ehab;Okasha, Hend;Samir, Safia;Nasr, Sami Mohamed;El-Kalamawy, Hadeer Adel;Saber, Mohamed Ali. And the article was included in Arabian Journal of Chemistry in 2022.Name: (2R,3R,4S,5R,6S)-2-(Hydroxymethyl)-6-(isopropylthio)tetrahydro-2H-pyran-3,4,5-triol The following contents are mentioned in the article:
Streptokinase (SK) is a potent plasminogen activator naturally produced by beta-hemolytic streptococcus bacteria and used as a thrombolytic drug. Optimize high yield production of recombinant streptokinase (rSK) in Escherichia coli and evaluate its thrombolytic activity. Synthetic gene encoding mature SK protein with optimization for rare codons and mRNA secondary structure was cloned into the expression vector pET-3a and transformed into Escherichia coli BL21 (DE3). Seed banks were established for high rSK expression clones. The native rSK protein expression was optimized using IPTG induction. The nonsol. rSK inclusion bodies were purified, denatured in 6 M guanidinium chloride, and refolded using the rapid dilution method. The refolded rSK protein was purified using anion exchange chromatog. and evaluated with ELISA. The activity of rSK was evaluated using the casein digestion method and in vitro blood clot lysis assay with reference drug Sedonase as standard Seed banks with high stable expression of native rSk (MW 47 kDa) were established. High rSK expression was optimized using 1 mM IPTG at bacterial OD600 0.6. The refolded rSK was prepared and purified successfully with high productivity (494 mg purified rsk/L culture). Using ELISA, the purified rSK mol. identity and conservation of native SK epitopes were confirmed. The enzymic activity of the purified rSK was 1.945×106 IU/mg with 62.94 ± 2.3% clot lysis efficiency. A high yield production of proper rSK protein with in vitro thrombolytic activity similar to com. SK has been achieved, suggesting a more cost-effective industrial production of its biosimilar drug. This study involved multiple reactions and reactants, such as (2R,3R,4S,5R,6S)-2-(Hydroxymethyl)-6-(isopropylthio)tetrahydro-2H-pyran-3,4,5-triol (cas: 367-93-1Name: (2R,3R,4S,5R,6S)-2-(Hydroxymethyl)-6-(isopropylthio)tetrahydro-2H-pyran-3,4,5-triol).
(2R,3R,4S,5R,6S)-2-(Hydroxymethyl)-6-(isopropylthio)tetrahydro-2H-pyran-3,4,5-triol (cas: 367-93-1) belongs to alcohols. Similar to water, an alcohol can be pictured as having an sp3 hybridized tetrahedral oxygen atom with nonbonding pairs of electrons occupying two of the four sp3 hybrid orbitals. Alcohols may be oxidized to give ketones, aldehydes, and carboxylic acids. These functional groups are useful for further reactions. Oxidation of organic compounds generally increases the number of bonds from carbon to oxygen (or another electronegative element, such as a halogen), and it may decrease the number of bonds to hydrogen.Name: (2R,3R,4S,5R,6S)-2-(Hydroxymethyl)-6-(isopropylthio)tetrahydro-2H-pyran-3,4,5-triol
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