• Journal of Microbiology and Biotechnology
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• v.16 no.2
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• pp.312-317
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• 2006

The chromosomal sprD gene encoding Streptomyces griseus protease D (SGPD), a chymotrypsin-like protease, was disrupted in Streptomyces griseus by insertion of the neomysin-resistance gene. The production of chymotrypsin activity of sprD disruptant was not completely abolished, but delayed by 24 h, compared with that of wild-type strain. The aerial mycelial formation of sprD disruptant was retarded, and specifically the formation of spores was not observed in the central region of colonies. However, normal morphological development into spores was observed in the marginal region of colonies. In addition, the production of yellow pigment that might be dependent on A-factor was also decreased in the sprD disruptant, compared with that of the wild-type strain. Introduction of the sprD gene, which was placed on a high copy-numbered plasmid into S. griseus ${\Delta}sprD$, partially restored the ability of morphological development, and a significant level of sporulation was observed. When the overexpression vector for sprD, pWHM3-D, was introduced in S. griseus, there was no significant change in the chymotrypsin activity or colonial morphology, in contrast to Streptomyces lividans, indicating the presence of a tight regulation system for the overexpression of the sprD gene in S. griseus.

• Journal of Microbiology and Biotechnology
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• v.22 no.6
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• pp.818-825
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• 2012

Keratinases are exciting keratin-degrading enzymes; however, there have been relatively few studies on their immobilization. A keratinolytic protease from Chryseobacterium sp. kr6 was purified and its partial sequence determined using mass spectrometry. No significant homology to other microbial peptides in the NCBI database was observed. Certain parameters for immobilization of the purified keratinase on chitosan beads were investigated. The production of the chitosan beads was optimized using factorial design and surface response techniques. The optimum chitosan bead production for protease immobilization was a 20 g/l chitosan solution in acetic acid [1.5% (v/v)], glutaraldehyde ranging from 34 g to 56 g/l, and an activation time between 6 and 10 h. Under these conditions, above 80% of the enzyme was immobilized on the support. The behavior of the keratinase loading on the chitosan beads surface was well described using the Langmuir model. The maximum capacity of the support ($q_m$) and dissociation constant ($K_d$) were estimated as 58.8 U/g and 0.245 U/ml, respectively. The thermal stability of the immobilized enzyme was also improved around 2-fold, when compared with that of the free enzyme, after 30 min at $65^{\circ}C$. In addition, the activity of the immobilized enzyme remained at 63.4% after it was reused five times. Thus, the immobilized enzyme exhibited an improved thermal stability and remained active after several uses.

• Journal of Microbiology and Biotechnology
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• v.27 no.2
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• pp.251-261
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• 2017

Initially, we screened 18 Aspergillus sojae-like strains from Aspergillus spp. isolated from meju (Korean traditional fermented soybean brick) according to their morphological characteristics. Because members of Aspergillus section Flavi are often incorrectly identified because of their phylogenetic similarity, we re-identified these strains at the morphological and molecular genetic levels. Fourteen strains were finally identified as A. sojae. The isolates produced protease and ${\alpha}-amylase$ with ranges of 2.66-10.64 and 21.53-106.73 unit/g-initial dry substrate (U/g-IDS), respectively, which were equivalent to those of the koji (starter mold) strains employed to produce Japanese soy sauce. Among the isolates and Japanese koji strains, strains SMF 127 and SMF 131 had the highest leucine aminopeptidase (LAP) activities at 6.00 and 6.06 U/g-IDS, respectively. LAP plays an important role in flavor development because of the production of low-molecular-weight peptides that affect the taste and decrease bitterness. SMF 127 and SMF 131 appeared to be non-aflatoxigenic because of a termination point mutation in aflR and the lack of the polyketide synthase gene found in other A. sojae strains. In addition, SMF 127 and SMF 131 were not cyclopiazonic acid (CPA) producers because of the deletion of maoA, dmaT, and pks/nrps, which are involved in CPA biosynthesis. Therefore, A. sojae strains such as SMF 127 and SMF 131, which have high protease and LAP activities and are free of safety issues, can be considered good starters for soybean fermentations, such as in the production of the Korean fermented soybean products meju, doenjang, and ganjang.

• Microbiology and Biotechnology Letters
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• v.13 no.4
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• pp.321-327
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• 1985

Cultivation conditions for the production of extracellular alkaline protease by a nonpiamentation Serratia sp. and purification of the enzyme were studied. The maximum enzyme level was obtained at the beginning of stationary phase when the organism was cultured on brain heart infusion medium at $25^{\circ}C$ under aeration (gyratory shaking, 180 cycles/min). The enzyme was purified about 100 fold with 16.5％ yield by ammonium sulfate precipitation, ammonium sulfate fractionation followed by DEAE-cellulose chromatography (1st and 2nd). The purified enzyme moved as a single symmetrical peak in the analytical ultracentrifuge. The enzyme demonstrated its maximum activity at pH 8.5-9.0 and 4$0^{\circ}C$ when vitamin-free casein was used as a substrate.

• Korean Journal of Microbiology
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• v.9 no.1
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• pp.32-38
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• 1971

In solid culture of Endomycopsis fibuliger No.55, Eudomuopsis javanensis No.112 and Candida tropicalis No.340, the conditions of enzyme (protease, anylase and cellulase) production and the influence of addition of $(NH_4)_2;SO_4$ were examined, and the results obtained were as follows. 10 Wheat bran medium is found to be the best on the enzyme production in case of simple material. The optimum conditions ; are water content added 100 to 120%, temperature 25 to $80^{\circ}C$ and incubation times 2 to 3 days. 2) The cellulase production was scarely produced in the case of Endomyopsis fibuliger No.55, as well as, the amylase production was scarely producted in the case of Endomycopsis javanensis No.112 and Candida tropicalis No.340. 3) The enzyme production was remarkably increased when 5% of$(NH_4)_2;SO_4$ as inorganic nitrogen sources was admixed to wheat bran. 4) When 5% of $(NH_4)_2;SO_4$ was admixed to medium, the ratio of protein increase was 10.2 to 17.7% in wheat bran medium and 10.6 to 17.9% in sweet potato cake medium.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.22 no.3
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• pp.138-146
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• 1989

This experiment was conducted to utilize the water soluble protein from the fish meal in wastewater as nitrogen source by alkaline protease producting bacteria and to investigate the culture condition of the production. G-12 and G-14 strains having the strong activity of the alkaline pretense were isolated from sea water. These strains were identified as Pseudomonas chlororaphis and Pseudomonas alcaligenes according to physiologycal characteristics, respectively. In enzyme production, galactose and casein for G-12 strain, and raffinose and the water soluble protein of the fish meal wastewater for G-14 strain was favorable as carbon and nitrogen source. An action of inhibition appeared in all of the metal salts used. The optimal temperature of enzyme production was $30^{\circ}C$ for all strains. Optimal initial pH for the enzyme formation in G-12 and G-14 strains was pH 10.0 and 8.0. When these two strains were incubated for $30\~35$ hours in the optimal production medium, the enzyme production reached at maximum.

• Journal of Microbiology and Biotechnology
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• v.23 no.6
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• pp.794-801
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• 2013

This paper reports the production and characterization of crude xylanase from the newly isolated Humicola sp. Ly01. The highest (41.8 U/ml) production of the crude xylanase was obtained under the optimized conditions (w/v): 0.5% wheat bran, 0.2% $KH_2PO_4$, and 0.5% peptone; initial pH 7.0; incubation time 72 h; $30^{\circ}C$; and 150 rpm. A considerable amount of the crude xylanase was induced using hulless barley bran or soybean meal as the carbon source, but a small amount of the enzyme was produced when supplementary urea was used as the nitrogen source to wheat bran. The crude xylanase showed apparent optimal cellulase-free xylanase activity at $60^{\circ}C$ and pH 6.0, more than 71.8% of the maximum xylanase activity in 3.0-30.0% (v/v) ethanol and more than 82.3% of the initial xylanase activity after incubation in 3.0-30.0% (v/v) ethanol at $30^{\circ}C$ for 2 h. The crude xylanase was moderately resistant to both acid and neutral protease digestion, and released 7.9 and 10.9 ${\mu}mol/ml$ reducing sugar from xylan in the simulated gastric and intestinal fluids, respectively. The xylooligosaccharides were the main products of the hydrolysis of xylan by the crude xylanase. These properties suggested the potential of the crude enzyme for being applied in the animal feed industry, xylooligosaccharides production, and high-alcohol conditions such as ethanol production and brewing.

• Korean Journal of Poultry Science
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• v.36 no.4
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• pp.343-350
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• 2009

This study was carried out to investigate the possibility to utilize feather meal by bacterial strains. A bacterial strain SE-103 producing keratinolytic enzyme was isolated from the soil of the poultry slaughterhouses. It was identified as Bacillus sp. by judging from its morphological and physiological characteristics. Subsequently the optimal culture conditions for the production of keratinolytic protease by Bacillus sp. SE-103 were investigated. The composition of optimal medium was 3.0% glucose, 0.4% urea, 0.2% $NaNO_3$, and 0.15% KCl. In addition, optimal initial pH and temperature were 6.0 and $35^{\circ}C$, respectively.

• Microbiology and Biotechnology Letters
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• v.28 no.6
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• pp.329-333
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• 2000

Alkaline bacterium producing a high pro-tease activity at low temperature was isolated by using enrichment culture from soil samples and identified as Bacil-lus sp. WRD-1 Cell growth was maximal at 10 hours and the optimal initial pH and culture time of culture condition for enzyme production was pH 7 and 10 hours, respectively. Temperature range of high enzyme activity were $10~40^{\circ}C$. The optimal pH and temperature for the enzyme activity were pH9 and $30^{\circ}C$.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.42 no.5
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• pp.461-467
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• 2009

A bacterial strain was screened and identified from sea sediments in Tongyeong coastal area in order to obtain proteases or peptidase cleaving C-terminal of glutamic acid. Strain TS0611, which showed the highest activity from 5 isolated protease producing strains, was selected and its properties investigated. Strain TS0611 was a gram-positive rod, $1.2\;{\mu}m$ in cell length, catalase positive, motility-positive, melanin-negative and grew at 15~$50^{\circ}C$, and hydrolyzed gelatin and casein. This strain was identified as Bacillus thuringiensis or Bacillus cereus based on results from the API system(API 50 CHB) which examined saccharides properties, fatty acid composition of cell wall, and 16S rRNA gene sequence.