• Proceedings of the Korean Society of Life Science Conference
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• 2001.02a
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• pp.36-36
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• 2001

• Journal of Life Science
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• v.23 no.2
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• pp.273-281
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• 2013

The purpose of this research was to investigate the production and characterization of alkaline protease from Micrococcus sp. PS-1 newly isolated from seawater. Micrococcus sp. PS-1 was grown in Luria-Bertani (LB) medium. Its optimal temperature and pH for growth were $30^{\circ}C$ and 7.0, respectively. The effect of nitrogen sources was investigated on optimal enzyme production. A high level of alkaline protease production occurred in LB broth containing 2% skimmed milk. The protease was purified in a 3-step procedure involving ultrafiltration, acetone precipitation, and dialysis. The procedure yielded a 16.43-purification fold, with a yield of 54.25%. SDS-PAGE showed that the enzyme had molecular weights of 35.0 and 37.5 kDa. Its maximum protease activity was exhibited at pH 9.0 and $37^{\circ}C$, and its activity was stable at pH 8.0-11.0 and $25-37^{\circ}C$. The protease activity was strongly inhibited by PMSF, EDTA, and EGTA. Taken together, the results demonstrate that the protease enzyme from Micrococcus sp. PS-1 probably belongs to a subclass of alkaline metallo-serine proteases.

• Mycobiology
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• v.31 no.4
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• pp.209-213
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• 2003

Aspergillus flavus K-03 isolated from poultry forming soil in Korea was studied for its ability to produce extracellular proteases on basal medium containing 2%(w/v) chicken feathers. The fungus was observed to be a potent producer of such enzymes. Keratinolytic enzyme secretion was the best at 15 days of incubation period at pH 9 and temperature $40^{\circ}C$. No relationship existed between the enzyme yield and increase of biomass. Enzyme production was suppressed by exogenous sugars in descending order arabinose>maltose>mannose>fructose. But glucose did not influence the enzyme activity. The keratinolytic enzyme released by the fungus demonstrated the ability to decompose keratin substrates as chicken feather when exogenous glucose was present. The keratinolytic activity was inhibited by $HgCl_2$ and serine-protease inhibitors such as phenymethylsulfonyl fluoride(100%), chymostain(88%), crystalline soybean trypsin inhibtor(80%), antipain(45%) and aprotinin(40%), and was not by cystein-protease and aspartyl-protease inhibitors. The enzyme activity is only partially inhibited by metallo-protease inhibitor. Thus, the enzyme secreted by A. flavus K-03 belongs to the alkaline serine-type protease.

• Journal of Microbiology and Biotechnology
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• v.10 no.1
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• pp.103-106
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• 2000

Bacillus cereus KCTC 3674 excretes at least two kinds of extracellular proteases into the growth medium. Two major bands of the protease activity with molecular weights of approximately 100 and 38 kDa were obtained after gelatin-SDS-PAGE. The protease with a molecular weight of 38kDa was identified as an extracellular neutral (metallo-) protease. The neutral protease was quite thermostabile but labile to alkaline pH. On the contrary, the 100-kDa protease was thermolabile but stable to alkaline pH. The production of 38-kDa neutral protease was strongly affected by temperature, oxygen, carbonylcyanied m-chlorophenylhydrazone(CCCP) that was defined as a protonophofre, and cerulenin which inhibited lipid synthesis and caused changes in the membrane composition. On the other hand, the production of the 100-kDa protease was strongly affected by only temperature and cerulenin. Quinacrine (0.2 mM), which inhibits the penicillinase-releasing proteases of Bacillus licheniformis, had no effect, whatsoever, on the production of extracellular proteases in B.cereus KCTC 3674.

• Microbiology and Biotechnology Letters
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• v.20 no.3
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• pp.295-301
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• 1992

An alkaline protease producing microorganism was isolated from soil and identified as Streptomyces griseus HC-1141. The optimum pH and temperature for the purified enzyme activity were 8.0 and $60^{\circ}C$, respectively. The enzyme was relatively stable in the pH range of 7.0-9.0 and at the temperature below $60^{\circ}C$. The activity of purified enzyme was inhibited by $Hg^{2+}$, $Cu^{2+}$, $Zn^{2+}$, $Ba^{2+}$ and $Fe^{2+}$, whereas activated by $Mn^{2+}$ and $Ca^{2+}$. $\varepsilon$-Amino caproic acid, 2,4-dinitrophenol and iodine did not show inhibitory effect on the activity of alkaline protease, but p-chloromercuribenzoic acid, ethylendiaminetetraacetic acid showed inhibitory effect on the enzyme activity. These result suggested that the protease was metalloenzyme, and require a reactive SH group for the activity. The reaction of this enzyme follows typical Michaelis-Menten kinetics with the $K_m$ value of $2.229{\times}10^{-4}$M and the $V_{max}$ of $46.08 {\mu}$g/min for casein. The activation energy for the alkaline protease calculated by Arrhenius equation was 3.643 kcal/mol. This enzyme hydrolyzed casein more rapidly than the hemoglobin and egg albumin.