• The Korean Journal of Mycology
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• v.14 no.4
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• pp.273-280
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• 1986

The degree of autolysis and lytic enzyme production in the culture filtrate of Rhizopus oryzae was investigated. The formation of protoplast by using autolytic enzymes from Rh. oryzae was also attempted. Protoplasts were liberated from Rh. oryzae mycelium by lytic enzymes present in autolytic-phase culture filtrate. Maximum release of chitosanase and proteolytic enzyme into culture filtrate during autolysis was corresponded to maximum protoplast-liberating activity. High yields of protoplasts were obtained from 10 hr-age of Rh. oryzae mycelium with 0.5 M mannitol as osmotic stabilizer. The optimum temperature and pH for mycelium digestion were $25{\sim}30^{\circ}C$ and $6.0{\sim}6.5$ respectively. The mycelium of the 18 hours cultures were treated with autolytic enzyme in same volume of osmotic stabilizer at $30^{\circ}C$ for 5 hours and then it was confirmed by scanning electoron microscope that protoplast were produced beside the digesting cell wall of the fungi.

• Microbiology and Biotechnology Letters
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• v.17 no.1
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• pp.69-73
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• 1989

Cellular autolytic enzyme was isolated from the supernatant fluid of exponentially growing cuiture of Cl. butyricum ID-113. The autolysin was partially pruified by ammonium sulfate fractionation, chromatography on DEAE-Sephadex A-50 and gel filtration through Sephadex G-200. This autolytic enzyme lysed SDS-treated cell wall fractions of Cl. butyricum ID, but not whole cells at all. Its optimum pH and temperature were 5.0 and 37$^{\circ}C$, respectively. This enzyme was relatively stable at neutral pH, but sensitive to heat treatment. Enzyme activity was not influenced by the addition of various divalent cation, but inhibited by Cu$^{++}$.

• Journal of the Korean Society of Food Science and Nutrition
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• v.30 no.5
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• pp.782-786
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• 2001

The taste characteristics and functionality of low-utilized small longfinned squid as affected by two stage enzyme hydrolysis were examined. In taste active-components, total free amino acid contents in hot-water and autolytic extract, two stage enzyme hydrolysate (TSEH) of longfinned squid were 2,792.5 mg%, 8,393.8 mg% and 9,186.1 mg%, respectively. The major free amino acids were Pro, Leu, Glu, Tau, Lys, Arg, Phe, Val and Ile. As for quarternary ammonium bases, betaine was the principal component (593.8 mg%) and also contents of TMAO, AMP in longfinned squid TSEH were 234.8% mg% and 51.0 mg%, respectively. The major inorganic ions in TSEH were Na(874.0 mg%), K (398.2 mg%), Cl (1,213.1 mg%) and PO$_4$(995.9 mg%). From the results in sensory tests, TSEH was superior to other extracts on the aspects of taste characteristics such as umami intensity, sweetness, taste harmony and transparency of extract. Also TSEH of longfinned squid revealed very higher Angiotensin-I converting enzyme inhibition ratio (92.1%) than those of hot-water and autolytic extract.

• Microbiology and Biotechnology Letters
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• v.17 no.1
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• pp.63-68
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• 1989

The optimum conditions for cellular autolysis in Clostridium butyricum ID-113 have been investigated. Cellular autolysis was optimal at pH 1.0 in 0.05 M potassium phosphate buffer and at 37$^{\circ}C$. The rate of cellular autolysis depended on the age of culture. The most rapid cellular autolysis occurred in the cells of mid-exponentially growing cultures, but cellular autolysis decreased sharply when the cultures entered the stationary phase. A growing culture of Cl. butyricum ID-113 was induced to autolyze and lost its turbidity spontaneously in the hypertonic NaCl, sucrose, or glucose medium. The autolytic enzyme activity was found In the autolysate of cells and the supernatant of the culture.

• Journal of Microbiology and Biotechnology
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• v.11 no.6
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• pp.957-963
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• 2001

Bacillus subtilis YL-1004 was isolated from soil for the development of agents to control dental caries. This strain produced an extracellular lytic enzyme that hydrolyzed the Streptococcus mutans cell wall. The lytic enzyme was purified to homogeneity by affinity chromatography and gel permeation chromatography to give a single band on SDS-PAGE and non-denaturing polyacrylamide gel electrophoresis. The molecular weight of the enzyme was deduced from SDS-PAGE and gel chromatography to be 38 kDa and the PI to be 4.3 from isoelectric focusing. Sirty $\%$ of its lytic activity remained after incubation at $50^{\circ}C$ for 30 min, and its optimal temperature was $37^{\circ}C$ . The enzyme showed its highest activity at pH 8.0 and was stable at pHs ranging from 4.0 to 9.0. Treatment with several modifiers showed that a cysteine residue was involved in the active site of the enzyme. This lytic enzyme from Bacillus subtilis YL-1004 exhibited specificity towards Streptococci and also showed autolytic activity on Bacillus subtilis YL-1004.

• Korean Journal of Food Science and Technology
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• v.30 no.6
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• pp.1345-1349
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• 1998

The flavor constituents and functionality of small kingfish were studied as affected by two stage enzyme hydrolysis (TSEH). Total free amino acid contents in water extract, autolytic extract and TSEH of small kingfish were 541.3 mg%, 8,245.3 mg% and 6,636.6 mg%, respectively. Major free amino acids in TSEH were hydroxyproline, glutamic acid, proline, leucine, phenylalanine, lysine, arginine. As for nucleotides and other bases, IMP, TMAO and total creatinine were principal components in TSEH. And the major inorganic ions in TSEH were Na, K, P, Cl and $PO_{4}$. Also TSEH of small kingfish revealed very higher Angiotensin-I converting enzyme inhibition effect than those of water and autolytic extract.

• Journal of Microbiology and Biotechnology
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• v.6 no.1
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• pp.1-6
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• 1996

A bacterial strain NS70 producing an alkaline protease was isolated from soil samples taken near a hot spring and identified as Bacillus licheniformis by its morphological and physiological properties and cellular fatty acid analysis. The isolated alkaline protease was purified by ammonium sulfate fractionation, DEAE-, CM-, and Phenyl-Sepharose column chromatography. The molecular weight of the purified enzyme was estimated to be 32, 000 Da by sodium dodecylsulfate polyacrylamide gel electrophoresis. Its optimal pH and temperature for proteolytic activity against Hammarsten casein were 12 and $65^{\circ}C$, respectively. The enzyme was stable at alkaline pH range from 6.0 to 12.0, and fairly stable up to $65^{\circ}C$. The enzyme was inhibited by phenylmethylsulfonyl fluoride but not by EDTA and N-ethylmaleimide indicating that the enzyme is serine protease. Enzyme activity was markedly inhibited by $Hg^{2+}$ and $Cu^{2+}$. Autolytic phenomena were observed on purified protease NS70 but autolysis was reduced by the addtion of $Ca^{2+}$ ion or bovine serum albumin.

• Bulletin of the Korean Chemical Society
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• v.10 no.6
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• pp.600-604
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• 1989

In order to find the rational methods for improving the thermal stability of subtilisin Carlsberg, the mechanisms of irreversible thermoinactivation of the enzyme were studied at $90^{\circ}C.$ At pH 4, the main process was hydrolysis of peptide bond. This process followed first order kinetics, yielding a rate constant of $1.26\;{\times}\;10^{-1}h^{-1}$. Hydrolysis of peptide bond of PMS-subtilisin occurred at various sites, which produced new distinct fragments of molecular weights of 27.2 KD, 25.9 KD, 25.0 KD, 22.3 KD, 19.0 KD, 17.6 KD, 16.5 KD, 15.7 KD, 15.0 KD, 13.7 KD, and 12.7 KD. Most of the new fragments originated from the acidic hydrolysis at the C-side of aspartic acid residues. However 25.0 KD, 15.7 KD, and 13.7 KD which could not be removed in purification steps stemmed from the autolytic cleavage of subtilisin. The minor process at pH 4 was deamidation at asparagine and/or glutamine residues and some extend of aggregation was also observed. However, the aggregation was main process at pH 7 with a first order kinetic constant of $16 h^{-1}.$ At pH 9, the main process seemed to be combination of deamidation and cleavage of peptide bond.

• Korean Journal of Food Science and Technology
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• v.24 no.6
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• pp.519-523
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• 1992

The patterns on the proteolysis of mussel protein using a commercial enzyme preparation were investigated. The best one among six commercial enzyme preparations for the manufacture of mussel extract was Corolase PP, based on the degree of hydrolysis (DH). When the raw mussel paste, without water addition, was adjusted to pH 6.5, added 0.1% (w/w dry basis) of Corolase PP. and reacted at $50^{\circ}C$ for four hours, it reached the maximum value of DH (79%). The precooking of raw mussel decreased the efficiency of extraction and hydrolysis of the protein, due to the inactivation of the autolytic enzymes contained in the mussel. During the course of proteolysis, major free amino acids such as glycine, alanine, glutamic acid and lysine, representing a characteristic brothy taste of mussel were replaced with free hydrophobic amino acids including valine, methionine, isoleucine, and leucine. The electrophoretic pattern and HPLC-GPC pattern of mussel protein hydrolysates during the hydrolysis were observed and also discussed.

• Korean Journal of Food Science and Technology
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• v.30 no.6
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• pp.1339-1344
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• 1998

To develop natural flavoring substances, optimal hydrolysis conditions for two stage enzyme hydrolysates (TSEH) using small kingfish (Maegari) were investigated. The optimal conditions for TSEH were revealed in temperature at $50^{\circ}C$ for 3 hours digestion with alcalase (Aroase AP-10, pH 8.0) at the 1st stage and 2 hours digestion at $45^{\circ}C$ with neutrase (Pandidase NP-2, pH 6.0) at the 2nd stage. From the results in quality tests of water extracts, autolytic extracts and 4 kinds of enzyme hydrolysates, TSEH processing method was superior to other methods on the aspects of yield, nitrogen contents, taste such as umami intensity and inhibition of off-flavor formation, and transparency of extracts. We may conclude that TSEH from small kingfish was more flavorable compared with the conventional seasoning materials, it could be utilized as the seasoning substances for fisheries processing.