• 한국미생물·생명공학회지
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• 제24권4호
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• pp.500-506
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• 1996

Bacillus subtilis SH-1 screened from coastal sea water of South Korea was used to produce bacteriolytic enzyme. The production of bacteriolytic enzyme by immobilized cells was investigated. The optimum conditions for the continuous production of the bacteriolytic enzyme using immobilized cells were 2.4 mm diameter of 0.3% alginate beads, 20 ml/h of substrate feeding rate and 20 l/min of aeration rate. A productivity of 76.5 to 88.0 units/ml could be obtained for 25 days by continuous column reactor under the optimum conditions.

• 한국미생물·생명공학회지
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• 제21권5호
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• pp.453-460
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• 1993

The extracellular bacteriolytic enzyme from alkalophilic Bacillus sp. YJ-451 was endopeptidase which hydrolyzes the peptide bond at the amino group of D-glutamic acid in the peptidoglycan. Protoplast transfomation system of B. subtilis by the lytic enzyme that differs, in mechanisms, from lysozyme which was used to transformation of B. subtilis was investigated. High protoplast yield was obtained from cells cultured in PAB at the late logarithmic growth phase.

• Journal of Microbiology and Biotechnology
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• 제3권2호
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• pp.73-77
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• 1993

The nucleotide sequence of Bacillus sp. bacteriolytic enzyme gene, lytP and its flanking regions were determined. A unique open reading frame for a protein of Mw. 27, 000, and a putative terminator sequence, were found behind a concensus ribosome binding site located 8 nt upstream from ATG start codon. The primary amino acid sequence deduced from nucleotide sequence revealed a putative protein of 255 amino acid residues with an Mw. of 27, 420. No significant homology could be found between the amino acid sequence of Bacillus sp. bacteriolytic enzyme and that of other cell wall hydrolases.

• 한국미생물·생명공학회지
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• 제23권5호
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• pp.580-587
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• 1995

In order to produce the bacteriolytic enzyme, bacterial strains capable of excreting a large amount of the enzyme were screened from the coastal sea water samples in Pusan. The bacterial strain SH-1, which showed the highest activity among 43 bacteriolytic enzyme producing bacteria, was finally selected for further studies. The strain SH-1 was an endospore-forming grampositive rod, and the position of spore was paracentral. These morphological characteristics assigned the isolated strain to the morphological group I classified by Gordon. The fatty acid composition of the bacterial stain was analyzed to be consisted of branched chains of iso-Cn and anteiso-Cn. Based on the percent content of the branched chain (93.85%), the isolates could be identified as a species of Bacillus. According to the experimental results of the API system (API 50CHB & API 20E) the strain was identified as Bacillus subtilis. Numerical texonomy, in which 82 major characters were examined using several species of Bacillus as the standard bacteria, indicated that the strain SH-1 showed 90% similarity to Bacillus subtilis. Thus, the isolated strain SH-1 could be identified as Bacillus subtilis.

• Journal of Microbiology and Biotechnology
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• 제2권3호
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• pp.161-165
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• 1992

The gene encoding the bacteriolytic enzyme cell wall peptidoglycan hydrolase from alkalophilic Bacillus sp. was cloned in E. coli using pBR322 as a vector. A recombinant plasmid, designated pYTR451, was isolated and the size of the cloned HindIII fragment was found to be 4.8 Kb. The cell wall hydrolysis activity of an extract of the E. coli harboring the recombinant plasmid pYTR 451 was detected by SDS- polyacrylamide gel containing 0.2% (w/v) purified cell wall of Bacillus sp. The molecular weight of the enzyme was estimated to be about 27, 000 corresponding to the molecular weight of the Bacillus sp. bacteriolytic enzyme. The recombinant plasmid was found to contain the fragment originated from Bacillus sp. YJ-451 chromosomal DNA by Southern hybridization.

• 한국식품위생안전성학회지
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• 제10권4호
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• pp.231-237
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• 1995

해양에서 분리한 용균활성이 우수한 균주인 Bacillus subtilis SH-1의 배양특성을 조사하여 다음과 같은 결과를 얻었다. Bacillus subtilis SH-1에 의한 용균효소 생산을 위한 치적조건인 1.0% glucose, 1.0% yeast extract, 1.0% NaCl, 0.02% $K_2HPO_4,\;0.002%\;MgSo_4{\cdot}7H_2O,\;0.001%\;MnSO_4{\cdot}5H_2O,\;0.0001%\;FeSO_4{\cdot}7H_2O\;(ph\;8.0)$의 배지로 $30^{\circ}C$에서 진탕배양하여 배양 28시간일 때 효소활성이 가장 높았다.

• Journal of Dairy Science and Biotechnology
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• 제21권2호
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• pp.114-119
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• 2003

Lactobacillus acidophilus CP4A 균주가 생산하는 acidocin 4A를 정제하고자 ammonium sulphate 침전법 , Octyl-sepharose CL-4B column chromatography, $C_{18}$ Sep-Pak cartridge, $C_{18}$ RP HPLC, HPLC gel filtration을 실시하였고 tricine SDS-PAGE를 통해 약 4.1 kDa의 박테리오신임을 확인하였다. Acidocin 4A의 항균작용 기작을 L. delbrueckii subsp. lactis ATCC 4797을 대상으로 TEM을 이용해 관찰한 결과 세포벽이 해리되고 세포벽사이로 세포내용물이 용출되어 궁극적으로 cell lysis가 일어나는 bacteriolytic 현상을 확인하였다. 또한, acidocin 4A의 생산에 관련된 유전자의 존재 위치를 파악하고자 EtBr을 이용한 curing방법을 실시하였으며 그 결과 약 20 kb 크기의 plasmid에 acidocin 4A 생산과 자체 면역에 관련된 유전자가 존재함을 알 수 있었다.

• 한국미생물·생명공학회지
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• 제24권2호
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• pp.191-196
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• 1996

The bacteriolytic enzyme produced from Bacillus subtilis SH-1 was purified and characterized, and its molecular weight was determined. The bacteriolytic enzyme activity was increased about 66.5 times via purification with recovery yield of 18.5%. The optimum pH and temperature of this enzyme were 9.0 and 50$\circ$C. The enzyme was stable within a pH range of 6.0-10.0 and unstable above 60 . The molecular weight of the enzyme was estimated to be 23,000 dalton in a form of monomer with no other subunits. Effect of the enzyme on the lysis of bacteria engaged in food posion was tested. The lysis degree was below 31% against Gram negative bacteria and above 48% in Gram positive bacteria. The values higher than 73% were obtained against Vibrio sp. and Listeria sp. As the turbidity of dissolved peptidoglycan clecreases, the free amino group levels were increased. And, based on hydrolysis of casein, this enzyme was thought to be an endopeptidase.

• Journal of Microbiology and Biotechnology
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• 제1권2호
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• pp.102-110
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• 1991

Alkalophilic Bacillus sp. YJ-451, which was isolated from soil at several area in Korea, produced a novel type of bacteriolytic enzyme (cell wall peptidoglycan hydrolase) extracellulary. The cell wall hydrolytic activity was identified as a clear zone on sodium dodecyl sulfate polyacrylamide gel electrophoresis containing 0.2% (w/v) cell wall of Bacillus sp. as substrate. This enzyme was successively purified 66 fold with 3.2% yield in culture broth by ammonium sulfate precipitation, CM-cellulose column chromatography, and gel filtration, followed by hydroxylapatite column chromatography. The molecular weight of the purified enzyme was estimated to be 27,000 by sodium dodecyl sulfate polyacrylamide gel electrophoresis and gel filtration column chromatography. The optimum pH and temperature for the activity of the enzyme were pH 10.0 and $50^{\circ}C$, respectively. The enzyme was stable between pH 5.0 and 10.0 and up to $40^{\circ}C$. Among the microorganisms used in this experiment the enzyme was active against most of gram negative strains and the genus Bacillus such as B. megaterium, B. licheniformis, B. circulans, B. pumilus, B. macerans, B. polymyxa. The release of dinitrophenylglutamic acid but not reducing group from cell wall peptidoglycan digested by the enzyme suggested that the enzyme is a kind of peptidase which hydrolyzes the peptide bond at the amino group of D-glutamic acid in the peptidoglycan.

• 한국미생물·생명공학회지
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• 제22권3호
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• pp.266-270
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• 1994

A bacteriocin was isolated from the supernatant fluid of M17G broth culture of Lactococcus sp. HY 449 strain, which showed strong inhibitory activity against the growth of selective indicator strain, Lactobacillus fermentum IFO3023. When the bacteriocin wasa added to the growing indicator cells or cell suspensions, viable cells and optical density were density were decreased, indicating bacteriolytic mode of action. Electron microscopic observation of indicator cells treated with bacteriocin revealed apparent damages on the cell surface and eventual lysis of cell walls.