• Korean Journal of Veterinary Research
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• v.39 no.1
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• pp.126-132
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• 1999

Swine plasminogen is not activated by staphylokinase of Staphylococcus aureus. In this study, the activation of swine plasminogen by staphylokinase of Staph hyicus subsp. hyicus was investigated and the effect of EDTA(disodium) on plasminogen activation was also studied. When the activation of swine plasminogen by staphylokinase of Staph hyicus subsp. hyicus was examined in fresh swine plasma, swine plasminogen could be weakly activated. However, when EDTA was added to the swine plasma, plasminogen activation was markedly enhanced, but this enhancement was not observed on bovine fibrin-dog plasminogen agar plate containing EDTA. Chicken and bovine plasminogens were not activated by staphylokinase of Staph hyicus subsp. hyicus. Using fresh swine plasma agar containing 0.07% EDTA, staphylokinase activity was detected in 96.3% of Staph hyicus subsp. hyicus strains isolated from pigs and in none of the chicken and bovine strains.

• Korean Journal of Veterinary Research
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• v.42 no.2
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• pp.201-208
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• 2002

Staphylokinase is seldom formed by animal Staphylococcus aureus strains. In this study, 76(72.4%) of 105 Staph aureus strains isolated from bovine mastitis were found to produce a staphylokinase. These staphylokinase producing strains were tested for lysogenic conversion by means of delysogenization procedure and isolation of serotype B and F converting phages. By the application of delysogenization method comprised of UV irradiation and acriflavine treatment to 76 staphylokinase-positive strains, the delysogenized cells could be observed in 29(38.2%) of the strains and delysogenization rates in 16(55.2%) of 29 delysogenized strains were 0.9% or less. A total of 7 serological group F phages were isolated from 76 staphylokinase-positive strains, and these phages could be again divided into three groups by the immunity reaction. Of 7 serotype F phages, 2 were isolated from the original lysogenic strains producing colonies of delysogenized cells after delysogenizing treatments and 5 were isolated from strains in which delysogenized cells were not observed after delysogenizing treatments. Difference of sensitivities to serological group F phages between original lysogenic strains and strains from which delysogenized cells were not isolated after delysogenizing treatments was not observed These data suggest that staphylokinase production of the remaining 42 strains might be also mediated by lysogenic conversion.

• Korean Journal of Veterinary Research
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• v.37 no.2
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• pp.359-365
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• 1997

Staphylococcus hyicus subsp. hyicus strains isolated from pigs, chickens and cattle were examined for the production of staphylokinase after inhibition of staphylococcal proteases by two procedures with EDTA(disodium). In one, EDTA was added to the bovine fibrin-dog plasminogen agar medium in concentration of 0.07% and paper strips soaked in 2mg/ml soy bean trypsin inhibitor were then applied on the agar plates. In the other, paper strips soaked in 5% EDTA solution were applied on the bovine fibrin-dog plasminogen agar plates and the strains to be tested were then streaked at right angles with the strip. By these procedures, staphylokinase activity was detected in 8(88.9%) of 9 strains from diseased pigs and in 57(80.3%) of 71 strains from skin of healthy pigs, but not in any strains from skin of healthy chickens and milk samples of mastitic cattle. Additionally kinase activity in 9 Staphylococcus species and subspecies isolated from bovine intramammary infections was also tested by these procedures. Staphylokinase activity was detected in 74.2% of Staph aureus strains and in 25% of Staph xylosus strains.

• Korean Journal of Veterinary Research
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• v.40 no.1
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• pp.49-55
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• 2000

Lysogenic conversion of Staphylococcus aureus to loss of ${\beta}-hemolysin$ production by serological group F phages is always associated with gain in staphylokinase production. In this study, the new phages belonging to serotype F were detected during the course of isolation of phages from Staph aureus of bovine origin and some characteristics of the new phages isolated were investigated. The new phages, ${\phi}470$ and ${\phi}499$, isolated from Staph aureus producing ${\beta}-hemolysin$ and staphylokinase(${\beta}^+\;K^+$) were found to convert ${\beta}^+\;K^+$ strain to ${\beta}^+K\;^+$, Staph aureus strains lysogenized by this serotype F single-converting phage ${\phi}470$ or ${\phi}499$ could be again lysogenized with serotype F double-converting phage ${\phi}506$. The frequency of lysogenization of indicator strains by serotype F single-converting phage was 100%, whereas the frequency for serotype F double-converting phage ${\phi}506$ varied from 4.2% to 97.6% according to the indicator strains. The indicator strain lysogenized with phage ${\phi}470$ was resistant to phage ${\phi}499$, and vice versa, but not to phage ${\phi}506$. Therefore, phage ${\phi}470$ and ${\phi}499$ were shown to be identical by immunity test.

• Biotechnology and Bioprocess Engineering:BBE
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• v.6 no.3
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• pp.167-172
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• 2001

Staphylokinase (SAK) was produced in B. subtilis using two different promoter systems, i.e. the P43 and sacB promoters. To maximize SAK expression in B. subtilis, fermentation control strategies for each promoter were examined. SAK, under P43, a vegetative promoter transcribed mainly by $\sigma$(sup)B containing RNA polymerase, was overexpressed at low dissolved oxygen (D.O.) levels, suggesting that the sigB operon is somewhat affected by the energy charge of the cells. The expression of SAK at the 10% D.O. level was three times higher than that at the 50% D.O. level. In the case of sacB, a sucrose-inducible promoter, sucrose feeding was used to control the induction period and induction strength. Since sucrose is hydrolyzed by two sucrose hydrolyzing enzymes in the cell and culture broth, the control strategy was based on replenishing the loss of sucrose in the culture. With continuous feeding of sucrose, WB700 (pSAKBQ), which contains the SAK gene under sacB promoter, yielded ca. 35% more SAK than the batch culture. These results present efficient promoter-dependent control strategies in B. subtilis host system for foreign protein expression.

• KSBB Journal
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• v.16 no.4
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• pp.415-419
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• 2001

To produce staphylokinase (SAK) in B. subtilis WB700, media optimization was carried out and the operation of batch and fed-batch fermentation were compared. Tryptone is a good nitrogen source and its optimum concentration in modified super rich(MSR) media is 15 g/L. When glucose is used as a limiting carbon source in the MSR media, 5 g/L of an optimum glucose concentration was identified for the SAK production under the control of P43 promoter. As the expression of P43 promoter is controlled by the limitation of oxygen, the SAK production was controlled at the 30% DO level in the fed-batch fermentation. Unexpectedly, batch fermentation using MSR media showed 1.5 times higher yield of SAK than that of the fed-batch fermentation. The main cause of the results comes from not achieving higher cell concentration in the fed-batch fermentation and the optimum expression level of P43 promoter under oxygen or nutrient limitations. We could not achieve the increase in cell concentration by any means in batch culture as well as fed-batch culture. The highest yield in the batch culture was 2880 units of SAK activity and 455 mg/L of secreted SAK.

• Journal of Microbiology and Biotechnology
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• v.22 no.7
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• pp.894-901
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• 2012

Based on their respective antitumor and thrombolytic activities, the superantigen staphylococcal enterotoxin C2 (SEC2) and staphylokinase (Sak) were chosen for the construction of the novel chimeric proteins Sak-linker-SEC2 and SEC2-linker-Sak using a linker composed of nine Ala residues. Both chimeric proteins possessed nearly the same PBMC proliferation stimulating activity and antitumor activity as SEC2 and thrombolytic activity as Sak. Neither the SEC2 or Sak component of each chimeric protein affected the activity of the other component. The results presented in this study provide a possible strategy to prevent and cure tumor thrombus.