• Microbiology and Biotechnology Letters
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• v.23 no.2
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• pp.145-149
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• 1995

For the purpose of developing a new biodegradable detergent, we have isolated a gene encoding wide-range temperature applicable alkaline protease from Xanthomonas sp. YL-37 (Lee et al., 1994, Kor. J. Appl. Microbiol. Biotechnol.). An alkaline protease gene was isolated from the gene bank that was prepared from the chromosomal DNA of Xanthomonas sp. YL-37. From the results of agarose gel electrophoresis and a restriction enzyme mapping, a 2.7 kb DNA fragment containing the alkaline protease gene was inserted in the plasmid pUC9. Extracellular activity of a clone having alkaline protease gene was detected on SDS-polyacrylamide gel with activity staining assay. The molecular weight of alkaline protease was determined to be about 64 kDa from 11% SDS-PAGE analysis. Alkaline protease activity, produced from E. coli which harboring the plasmid, showed no difference at reaction temperature 20, 30 and 40$\circ$C, respectively. This result showed that alkaline protease produced from E. coli harboring the plasmid was apparently the same as that of Xanthomonas sp. YL-37.

• KSBB Journal
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• v.5 no.4
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• pp.411-414
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• 1990

The extraction behavior of an alkaline protease using reversed micelles was investgated. The reversed micellar solution consisted of AOT in isooctane. It was found that distribution of arkaline protease into the organic phase increased at lower pH, lower ionic strength, and higher AOT concentration. When the real fermentation broth was extracted of alkaline protease, an activity yield of 20% and a purification factor of 2.0 were obtained.

• Microbiology and Biotechnology Letters
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• v.8 no.2
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• pp.77-85
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• 1980

This experiment was conducted to investigate the conditions for production and the characteristics of pretenses. Aspergillus oryzae KC-15, which is selected as a superior strain for the production of the protease, was used in this study. The results obtained were as follows: 1. The optimum culture time for the production of acid, neutral and alkaline protease on wheat bran medium were about 48, 48 and 72hr, respectively. The protease-produced by the strain were mainly alkaline and neutral one, but the production of acid protease was feeble extremely. 2. The addition of NaH$_2$PO$_4$, Na$_2$HPO$_4$, glucose, rice powder and Na-glutamate respectively to wheat bran media were effective for the production of alkaline and neutral protease, and the addition of (NH$_4$)$_2$HPO$_4$, glucose and rice powder respectively were effective for the production of acid protease. 3. Characteristics of professes(equation omitted) 4. As a heat resistance agent, NaH$_2$PO$_4$was the most effective one. The optimum amount of NaH$_2$PO$_4$was 10mg for alkaline and neutral protease, and 5mg for acid protease. 5. The heat resistance of the Protease by NaH$_2$PO$_4$was not recognized mostly above 6$0^{\circ}C$. 6. After the treatment of enzyme solution with 10mg of NaH$_2$PO$_4$for 30 minutes at 55$^{\circ}C$, the residual activities measured for alkaline, neutral and acid protease were 58, 57 and 55％ respectively.

• Korean journal of applied entomology
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• v.27 no.4
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• pp.211-218
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• 1988

Polyhedral proteins and the endogenous alkaline protease associated with larval-derived polyhedra of nuclear viruses isolated from Spodoptera litura, Bombyx mori, and Hyphantria cunea were investigated. Polyhedral proteins prepared under alkaline protease heat-inactivated condition were separated as one band with 31Kd in S. litura a H. cunea NpV and 30Kd in B. mori NPV by the SDS-polyacrylamide gel electroptoresis. Whereas polyhedral proteins without heat-inactivation were degraded into smaller polypeptides with a certain pattern in alkaline solution. The results of double-immunodiffusion and western blot analysis with antisera against polyhedral proteins indicated that those three polyhedral proteins had common antigenic determinants and the degradation of polyhedral proteins by alkaline protease could be confirmed.

• Microbiology and Biotechnology Letters
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• v.19 no.1
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• pp.1-7
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• 1991

A bacterial strain No.71, which produced alkaline protease, was isolated from soil and identified to the genus Bacillus. With the successive mutation, a mutant strain No. M-71, having high alkaline protease productivity, was obtanined from the parental strain No 71. Alkaline protease productivity of mutant strain No. M-71 was about 50 times as much as that of the parental strain No.71. The enzyme preparations showed strong activities toward casein, the optimum pH being 11.0 and the optimum temperature about $55^{\circ}C$.

• Applied Biological Chemistry
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• v.31 no.4
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• pp.356-360
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• 1988

The alkaline protease producing bacteria isolated from soil and identified as Bacillus subtilis. The optimum medium for alkaline protease production from the microorganism was as follows; soluble starch, 1.5% ; proteose peptone, 0.5% ; $K_2HPO_4$, 0.1% ; $MgSO_4{\cdot}7H_2O$, 0.02% and sodium carbonate, 1.0%. The optimum temperature for alkaline protease production was $35^{\circ}C$, and the initial pH of medium was pH 10.5. The alkaline protease activity was about 2,300 U per ml of culture broth by Casein-Folin Method. A 9.2 fold purification of alkaline protease was obtained from culture broth. The recovery was 14% and purified enzyme was identified as single band, and its molecular weight was about 19,000. The optimum temperature for enzyme reaction was $70^{\circ}C$, and optimum pH was 12. The activity of purified enzyme was inhibited by metal ion ($Fe^{++}$), and Phenylmethylsulfonyl Fluoride, a serine protease inhibitor.

• Journal of Microbiology and Biotechnology
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• v.19 no.4
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• pp.378-386
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• 2009

A sequential optimization strategy, based on statistical experimental designs, is employed to enhance the production of alkaline protease by a Bacillus pseudofirmus local isolate. To screen the bioprocess parameters significantly influencing the alkaline protease activity, a 2-level Plackett-Burman design was applied. Among 15 variables tested, the pH, peptone, and incubation time were selected based on their high positive significant effect on the protease activity. A near-optimum medium formulation was then obtained that increased the protease yield by more than 5-fold. Thereafter, the response surface methodology(RSM) was adopted to acquire the best process conditions among the selected variables, where a 3-level Box-Behnken design was utilized to create a polynomial quadratic model correlating the relationship between the three variables and the protease activity. The optimal combination of the major medium constituents for alkaline protease production, evaluated using the nonlinear optimization algorithm of EXCEL-Solver, was as follows: pH of 9.5, 2% peptone, and incubation time of 60 h. The predicted optimum alkaline protease activity was 3,213 U/ml/min, which was 6.4 times the activity with the basal medium.

• Food Science and Preservation
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• v.11 no.2
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• pp.227-232
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• 2004

• Microbiology and Biotechnology Letters
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• v.18 no.4
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• pp.344-351
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• 1990

An alkalophilic microoganism producing a detergent-resistant alkaline protease was isolated from soil and identified as Baeiltus sp. The alkaline protease has been partially purified by ammonium sulfate fractionation, DEAE-Cellulose, CM-Cellulose and Sephdex G-100 column chromatography. The purified alkaline protease was highly active at pH 12-13 toward casein and stable at pH values from 6 to ll. The optimum temperature for the enzyme reaction was $55^{\circ}C$. The enzyme was completely inactivated by diisopropyl fluorophosphate (DFP) indicating that the enzyme was serine protease, but considerabiy stable in the presence of surface active agents.

• Applied Biological Chemistry
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• v.43 no.3
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• pp.169-173
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• 2000

For the development of enzyme detergent capable of effectively washing at low temperature, a bacterium producing alkaline protease was isolated from soil samples, and properties of the enzyme were investigated. The selected strain was Gram negative, rod shape$(0.6{\sim}0.7{\times}1.3{\sim}2.6\;{\mu}m\;in\;size)$ and motile. It had the degradation activity of aesculin, gelatin and casein, and was catalase-positive. The cell wall components was meso-DAP, and G+C mole contents was 43.3%. From these results, the strain was identified as Acinetobacter sp. KN-27. The activity of alkaline protease by this strain peaked with 3,300 D.U/mL after 36 hours in the liquid culture at $40^{\circ}C$. The optimal pH and temperature of the enzyme were pH 9 and $60^{\circ}C$, respectively. Alkaline protease produced by Acinetobacter sp. KN-27 has shown two active bands on the electrophoresis of native gel.