• 미생물학회지
• /
• 제28권2호
• /
• pp.162-168
• /
• 1990

토양에서 분리한 방사균 주에서 호염기성 단백질 분해효소의 생합성과 세포분화와의 관계를 규명하고자 기균사와 포자의 형성, 그리고 단백질 분해효소의 생산에 대한 배양조건의 영향을 조사하였다. 그 결과, 기질의 농도가 단백질 분해효소, 포자, 그리고 기균사의 형성의 조절에 매우 중요하며 이것은 배양액의 pH가 산성으로 변화기 문임을 알았다. 인산염완충용액을 이용하여 배양액의 pH를 6으로부터 9로 조정하여 주었을 때 단백질 분해효소의 생성은 pH가 증가함에 따라 증가하였다. 이와 같은 결과로부터 배양액의 pH가 호염기성 단백질 분해효소 생합성의 조절에 중요한 요소로 작용한다고 판단하였다. 판단하였다.

• 미생물학회지
• /
• 제24권1호
• /
• pp.32-37
• /
• 1986

In fermentation studies it revealed that Streptomyces sp. SMF 3001 started to synthesize extracellular alkaline protease from early exponential phase of cell growth. The biosynthesis of the alkaline protease was greatly induced by skim milk as a sola nitrogen source and further stimulation was observed under inorganic sulphur limited culture. However, it was found that the biosynthesis was apparently repressed by $NH_4^+$ and free amino acids, specially by cysteine. It was considered that the strain SMF 301 of Streptomyces sp. would produce the alkaline protease for the uptake of sulphur compounds from protein contained in the culture broth.

• Journal of Microbiology
• /
• 제33권2호
• /
• pp.115-119
• /
• 1995

The alkaline protease of Xanthomonas sp. YL-37 has been purified, and the properties of the enzyme investigated. The alkaline protease of Xanthomonas sp. YL-37 was purified form crude enzyme by ammonium sulfate fractionation, CM-cellulose ion exchange chromatography, and Sephadex G-100 gel filtration. Through the series of chromatographies, the enzyme was purified to homogenecity with specific activity of 4.23 fold higher than that of the crude broth. The molecular weight of the purified protease has been estimated to be 62 KDa on SDS-polyacrylamide gel electrophoresis. The optimal pH and temperature for alkaline protease activity were 11.0 and 50.deg.C, respectively. The enzyme was stable between pH 5.0 and 10.0 and up to 50.deg.C. Enzyme activity was lost up to 50% on heating at 70.deg.C for 30 minutes. The activity of alkaline protease was inhibited by Cu$\^$2+/, Zn$\^$2+/, Hg$\^$2+/, PMSF, and activated by Mn$\^$2+/ and Ca$\^$2+/. The $K_{m}$ value for casein as a substrate was 4.0 mg/ml.

• Journal of Microbiology and Biotechnology
• /
• 제33권2호
• /
• pp.195-202
• /
• 2023

Protease is a widely used enzyme particularly in the detergent industry. In this research, we aimed to isolate alkaline protease-producing bacteria for characterization as a laundry detergent additive. The screening of alkaline protease production was investigated on basal medium agar plus 1% skim milk at pH 11, with incubation at 30℃. The highest alkaline protease-producing bacterium was 6BS15-4 strain, identified as Bacillus gibsonii by 16S rRNA gene sequencing. While the optimum pH was 12.0, the strain was stable at pH range 7.0-12.0 when incubated at 45℃ for 60 min. The alkaline protease produced by B. gibsonii 6BS15-4 using dairy effluent was characterized. The optimum temperature was 60℃ and the enzyme was stable at 55℃ when incubated at pH 11.0 for 60 min. Metal ions K⁺, Mg²⁺, Cu²⁺, Na⁺, and Zn²⁺ exhibited a slightly stimulatory effect on enzyme activity. The enzyme retained over 80% of its activity in the presence of Ca²⁺, Ba²⁺, and Mn²⁺. Thiol reagent and ethylenediaminetetraacetic acid did not inhibit the enzyme activity, whereas phenylmethylsulfonyl fluoride significantly inhibited the protease activity. The alkaline protease from B. gibsonii 6BS15-4 demonstrated efficiency in blood stain removal and could therefore be used as a detergent additive, with potential for various other industrial applications.

• Journal of Microbiology and Biotechnology
• /
• 제21권12호
• /
• pp.1250-1256
• /
• 2011

In this study, Bacillus licheniformis cells were immobilized by entrapment in calcium alginate beads and were used for production of alkaline protease by repeated batch process. In order to increase the stability of the beads, the immobilization procedure was optimized by statistical full factorial method, by which three factors including alginate type, calcium chloride concentration, and agitation speed were studied. Optimization of the enzyme production medium, by the Taguchi method, was also studied. The obtained results showed that optimization of the cell immobilization procedure and medium constituents significantly enhanced the production of alkaline protease. In comparison with the free-cell culture in pre-optimized medium, about 7.3-fold higher productivity was resulted after optimization of the overall procedure. Repeated batch mode of operation, using optimized conditions, resulted in continuous production of the alkaline protease for 13 batches in 19 days.

• 한국미생물·생명공학회지
• /
• 제6권1호
• /
• pp.9-16
• /
• 1978

인삼액기스의 첨가량을 달리한 배지에 시간별로 국균의 첨가력($\alpha$-amylase, $\beta$-amylase, protease)에 미치는 영향을 조사한 결과는 다음과 같다. 1) Asp. oryzae의 $\alpha$-amylase 활성도는 48시간 배양까지는 대조구보다 다소 높았으나 그 이후는 같은 경향을 나타내었고, Asp. oryzae의 $\beta$-amylase 활성도는 배양 24시간에서 인삼엑기스, 첨가량순으로 촉진되어 1%구까지 증가하고 3~5%구는 그 활성이 첨가량 순으로 감소되었다. Asp. oryzae의 protease 활성도에서 acid protease는 배양 72시간까지 대조구보다 활성이 높았고, 그중 3.0%구의 48시간때가 가장 역가가 높았으며 120시간 배양에서는 인삼엑기스 3.0% 이상이 효소활성의 감소가 컸다. Alkaline protease는 대조구보다 객처리구의 역가가 떨어졌다. 그러나 alkaline protease는 aeid protease보다 활성도가 높았다. 2)Asp. niger의 $\alpha$-amylase 활성도는 인삼엑기스 첨가량 순으로 높아지고 그 활성이 급증가하였다가 급감소되는 경향을 보였고, Asp. niger의 $\beta$-amylase 활성도는 인삼엑기스, 첨가량에 따라 활성이 높아졌으며 0.5~3.0% 첨가시에는 대조구 및 5.0%보다 활성이 빨고 높게 나타났으며 5.0구는 그 활성이 억제되었다. Asp. niger의 protease 활성도는 acid protease가 alkaline protease보다 효소역가가 현저하게 높았다. acid protease는 배양 48시간까지는 인삼엑기스의 첨가량에 따라 효소역가가 조해되었으나 72시간에서는 인삼엑기스 첨가량 순으로 역가가 높았고, 단 5.0%구만 현저하게 효소력이 감소되었다. Alkaline protease는 배양 24시간에서 48시간까지는 0.1%구와 0.5%구가 대조구보다 효소력이 높았고 그 이상의 인삼엑기스가 첨가된 처리구보다 효소력이 낮았다. 그러나 48시간 이후부터는 효소력은 인삼엑기스 첨가량 순으로 감소를 나타내었다.

• Journal of Microbiology and Biotechnology
• /
• 제11권4호
• /
• pp.558-563
• /
• 2001

A new alkalophilic strain of Bacillus pumilus JB¬05 producing bleach-stable and halo-tolerant alkaline protease was isolated from cement industry effluents in Karnataka, India. The effects of carbon and nitrogen sources on protease production by this alkalophilic strain were observed after a 30-h incubation. A high level of alkaline protease activity was obtained in the presence of starch as the carbon and peptone as the nitrogen sources. The partially purified enzyme showed an optimum temperature and pH activity at $58^{\circ}C$ and 10.5, respectively. The enzyme was completely inhibited by PMSF (95.0%) indicating it as a serine protease. It is bleach-stable as it retained 35% original activity in the presence of 10% (v/v) hydrogen peroxide at $30^{\circ}$C after 2 h and is halo-tolerant as it retained 70% original activity in the presence of 2.5 M sodium chloride at $30^{\circ}C$ after 2 h incubation.

• Mycobiology
• /
• 제32권2호
• /
• pp.74-78
• /
• 2004

An alkaline protease produced by Aspergillus niger C-15 was purified and characterized. The enzyme was purified 19.41-fold with a specific activity of 74150 U/mg and a recovery of 34.4% by gel filtration and ion exchange chromatography. The molecular weight of the enzyme was estimated to be 34 kDa. The optimum pH and temperature for the protease activity were pH 8.0 and $60^{\circ}C$, respectively. The enzyme activity inhibited by EDTA suggests that the preparation contains a metalloprotease. The enzyme activity of the metalloprotease was completely inhibited by 5 mM $HgCl_2$ and $FeCl_3$, while partially inhibited by $CuSO_4$, and $MnCl_2$. When polyols such as glycerol, mannitol, sorbitol and xylitol, were added to the reaction medium, most polyols tested enhanced protease activity. Especially, glycerol showed the highest effect. The alkaline metalloprotease was stable at high temperature and retained more than 90% of the initial activity at $60^{\circ}C$ and 86.4% under addition of glycerol.

• 한국미생물·생명공학회지
• /
• 제17권4호
• /
• pp.358-364
• /
• 1989

토양으로부터 분리한 Streptomyces sp. YSA-130으로부터 활성이 좋은 결정화된 alkaline protease를 분리하였다. Alkaline protease 생산의 최적 배양조건은 2.0% soluble starch, 1.0% soytone, 0.3% $K_2$HPO$_4$, 0.02% MgSO$_4$.7$H_2O$, 0.8% $Na_2$CO$_3$ 3$0^{\circ}C$, pH 10.5에서 72 시간 배양하였을 때 였다. Alkaline protease이 정제는 (NH$_4$)$_2$SO$_4$. 분별침전, 투석, DEAE cellulose column chromatography, Sephadex G-75 gel filtration, crystallization으로 하였으며, 그 결과 비활성도 14,290unit/mg, 정제도 23.8 배였고, 수율은 20.0% 이었다. Alkaline protease의 반응 최적온도와 pH는 6$0^{\circ}C$ 와 11.5이었으며, 효소의 pH 안정성은 5.5-12.0에서 안정하였고, 온도 안정성은 5$0^{\circ}C$까지 안정하였으며, $Ca^{++}$ ion 첨가시 6$0^{\circ}C$까지 안정성이 증가하였다. Alkaline protease의 분자량은 30,000이었으며 금속이온, EDTA, 환원제는 활성에 영향이 없었고 DFP에 의해 저해되었다. 계면활성제에 저항성이 크고 $H_2O$$_2$에 대한 잔존활성은 60% 을 유지하였다.

• 한국미생물·생명공학회지
• /
• 제6권1호
• /
• pp.33-40
• /
• 1978

Immobilization of alkaline protease was investigated by absorbing the enzyme on adsorbents. Alkaline protease was adsorbed on silica gel selected as a carrier to immobilize the enzyme. In this study, properties of the immobilized enzyme were compared with those of the soluble enzyme. 1) The optimum pH (10.0) of the enzyme was not changed, but the activity was increased at alkaline pH by immobilization. 2) The optimum temperature of the immobilized enzyme was shifted from 50$^{\circ}C$ to 45$^{\circ}C$, while the temperature-activity Profile became broader than those of the soluble enzyme. 3) The pH stability of the immobilized enzyme was significantely increased at pH 4.0, althouth it did not change in the neutral and alkaline pH region. 4) The heat stability of the enzyme was enhanced in the temperature range of 55$^{\circ}C$∼65$^{\circ}C$ by the immobilization. 5) The immobilized enzyme retained 40％ of its original activity after repetitive use for 6 times. 6) The enzyme stability was greately improved for a prolonged storage at 4$^{\circ}C$.