Soybean-milk-coagulating activity of Bacillus pumilus derives from a serine proteinase

Aoyama, Misako; Yasuda, Masaaki; Nakachi, Koh; Kobamoto, Naotada; Oku, Hirosuke; Kato, Fumio

doi:10.1007/s002530051631

Cited by 35 publications

(25 citation statements)

References 13 publications

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“…The optimum temperature for the activity of the purified protease was 40°C with 50% relative activity at 50°C (Figure 3). This is really unusual and surprising since some previous reports on proteases from Bacillus species have temperature optima of 50°C to 60°C [5,[15][16][17][18][19][20][21]. The result obtained from the present study is however similar to those of reports by Feng et al [22] and Park and Cho [23] on proteases from Bacillus pumilus strain and Bacillus sp.…”

Section: Purification Of Extracellular Protease From B Brevis Mwb-01supporting

confidence: 85%

Purification and partial characterization of serine alkaline metalloprotease from Bacillus brevis MWB-01

Olajuyigbe

Falade

2014

Bioresour. Bioprocess.

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Background: Proteases from bacteria are among the most important hydrolytic enzymes that have been studied due to their extracellular nature and high yield of production. Of these, alkaline proteases have potential for application in detergent, leather, food, and pharmaceutical industries. However, their usefulness in industry is limited by low activity and stability at high temperatures, extreme pH, presence of organic solvents and detergent ingredients. It is therefore very crucial to search for new alkaline proteases with novel properties from a variety of microbial sources. Results: In the present study, 21 Bacillus species isolated from organic waste sites were screened for proteolytic activity on casein agar. Bacillus brevis MWB-01 exhibited highest proteolytic activity with a clear zone diameter of 35 mm. Production of protease from B. brevis MWB-01 was investigated in optimized media after 48 h of cultivation with shaking (180 rpm) at 37°C. The protease was partially purified in a two-step procedure using ammonium sulphate precipitation and gel filtration chromatography on Sephadex G-200 column. The enzyme was purified 2.1-fold with yield of 4.6%. The purified protease had optimum temperature of 40°C with relative activity of about 50% at 50°C and was uniquely stable up to 60°C after 30 min of incubation exhibiting 63% residual activity. The enzyme had optimum pH of 8.0 and remarkably showed relative activity above 70% at pH 9.0 to 11.0 and 53% at pH 12.0, respectively and was very stable over a wide pH range (6.0 to 12.0). Ca 2+ and Mn 2+ increased protease activity with 9.8% and 3.5%, respectively; Hg 2+ and Zn 2+ strongly inhibited protease activity by 89% and 86%. The almost complete inhibition of the enzyme by phenylmethylsulphonyl fluoride (PMSF) and ethylene diamine tetra acetic acid (EDTA) confirmed the enzyme as a serine metalloprotease. The enzyme had highest compatibility with Sunlight, a commercial laundry detergent.

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Section: Purification Of Extracellular Protease From B Brevis Mwb-01supporting

confidence: 85%

Purification and partial characterization of serine alkaline metalloprotease from Bacillus brevis MWB-01

Olajuyigbe

Falade

2014

Bioresour. Bioprocess.

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“…The enzyme was completely inhibited by 1 mM DFP, a well-known inhibitor of serine protease 4,21 suggesting that this enzyme was a serine protease. Similar results of serine proteases completely inhibited by DFP were observed in serine protease produced by B. licheniformis 22 , B. pumilus 7,21 and B. intermedius 3-19 23 . the enzyme under denaturing condition was estimated to be 27 kDa, similar to the molecular masses of alkaline proteases, which were in the ranges of 15 to 30 kDa 11 .…”

Section: Inhibitors Of the Protease Inhibitors Of The Protease Inhibisupporting

confidence: 73%

Untitled

Yossan¹,

Reungsang²,

Yasuda³

2006

ScienceAsia

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An alkaline protease produced by Bacillus megaterium isolated from a fermented broth of Thai fish sauce was purified by hydrophobic interaction combined with gel filtration techniques. After final purification step, the enzyme was purified 148-fold with an increase in specific activity from 0.09 to 13.33 U/mg protein.The properties of the purified enzyme were then analyzed. The molecular weight of the enzyme under denaturing condition was estimated to be 27 kDa. The optimum pH and temperature for protease activity were 10 and 50 o C, respectively. This protease could retain the activity in the pH and temperature ranging from pH 7.5 to 9.5 and 30 to 45 o C, respectively, resulting in the relative activity of higher than 80%. The enzyme was completely inhibited by diisopropyl fluorophosphate (DFP) suggesting that it was a serine protease. Phenylmethylsulfonyl fluoride (PMSF), p-Chloromercuribenzoic acid (PCMB), ethylenediaminetetraacetic acid (EDTA), 1, 10-phenanthroline and Fe 3+ strongly inhibited the activity of this purified enzyme activity, decreasing relative activity to lower than 15%. Protease activity was enhanced by Mn 2+ , Ca 2+ and Mg 2+ . Cytochrome C, soybean protein isolate and casein were good substrates specific to the enzyme with the relative activity of more than 100%.

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“…The pH stability of the alkaline protease from B. subtilis CN2 was reported to be in the range of 7-11 (Uchida et al, 2004). The optimum temperature for Bacillus pumilus (Aoyama et al, 2000) and B. subtilis CN2 protease (Uchida et al, 2004) was 50 8C which was similar to our parental protease. However, a higher optimum temperature of 60 8C was observed in Bacillus sp.…”

Section: Fungimentioning

confidence: 65%

Fungal protease: Production, purification and compatibility with laundry detergents and their wash performance

Savitha

Sadhasivam

Swaminathan

et al. 2011

Journal of the Taiwan Institute of Chemical Engineers

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Soybean-milk-coagulating activity of Bacillus pumilus derives from a serine proteinase

Cited by 35 publications

References 13 publications

Purification and partial characterization of serine alkaline metalloprotease from Bacillus brevis MWB-01

Purification and partial characterization of serine alkaline metalloprotease from Bacillus brevis MWB-01

Untitled

Fungal protease: Production, purification and compatibility with laundry detergents and their wash performance

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