• 한국미생물·생명공학회지
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• 제13권3호
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• pp.279-283
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• 1985

L-Lysine생산균주 육종의 한 방법으로, Brevibacterium flavum과 Corynebacterium glutamicum의 이속간 원형질체 융합을 실시하였으며, 이들 균주에 대한 원형질체 형성과 재생의 최적 조건을 조사하였다. 그 결과, Corynebacterium glutamicum ATCC 21514 S의 경우, lysozyme을 300$\mu\textrm{g}$/$m\ell$의 농도로 처리하였을 때 12시간 경과 후 99%의 원형질체 형성과 12%의 재생율을 보였으며, Brevibacterium flavum ATCC 21528R 은 lysozyme을 400$\mu\textrm{g}$/$m\ell$로 처리했을 때 12시간 경과 후 99%의 원형질체 형성과 10%의 재생율을 보였다. Brevibacterium flavum ATCC 21514 S 의 이속간 원형질체 융합에서 PEG 농도별 실험을 하여본 결과 PEG 6,000, 30% (w/v)를 사용함으로써 재생세포당 1.2$\times$$10^{-5}$의 재조합 빈도를 얻었으며, 여기에서 얻어진 재조합주들 가운데 KR$_{43}$ 주는 L-lysine 생성능이 모균보다 12% 증가를 나타내었으며, as-partokinase 효소 활성 측정치는 모균보다 13% 높은 것으로 나타났다.

• 한국미생물·생명공학회지
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• 제15권2호
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• pp.104-111
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• 1987

Lysine 생산균주를 개량하기 위한 시도로서 Brevibacterium flavum ATCC 21528R 과 Brevibacterium flvum ATCC 21529S의 동종간 및 Brevibacterium flavum ATCC 21528R 과 Corynebacterium glutamicum ATCC 13058S 와 의 이속간 원형질체 융합을 실시하였다. 이들 균주들에 대한 원형질체 형성의 최적조건을 조사하고 재생과 융합에서의 plasma expander의 효과를 검토하였다. 융합주 No. CH23과 No. CH41은 최적 배양조건 하에서 L-lysine 생산성이 모균에서보다 각각 21％와 8.9％ 향상된 것이었다. L-Lysine 생합성 회로상의 중심효소인 asparto-kinase를 포함한 주요효소의 활성을 측정하였고, 융합주 No. CH23과 No. CH41에서의 L-lysine 생합성 대사제어를 친주와 비교하였다.

• Biotechnology and Bioprocess Engineering:BBE
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• 제10권3호
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• pp.225-229
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• 2005

To investigate impediments to plasmid transformation in Brevibacterium flavum BF4 and B. lactofermentum BL1, cell surface barriers were determined by measuring growth inhibition whilst enzymatic barriers were determined by comparing DNA methylation properties. B. lactofermentum was more sensitive to growth inhibition by glycine than B. flavum. Release of cellular proteins during sonication was more rapid for B. lactofermentum than for B. flavum. Plasmid DNA (pCSL 17) isolated from B. flavum transformed recipient $McrBC^+$ strains of Escherichia coli with lower efficiency than $McrBC^-$. McrBC digestion of this DNA confirmed that B. flavum contain methylated cytidines in the target sequence of McrBc sequences but B. lactofermentum contained a different methylation pattern. DNA derived from the B. lactofermentum transformed recipient $EcoKR^+$ strains of E. coli with lower efficiency than $EcoKR^-$, indicating the presence of methylated adenosines in the target sequence of EcoK sequences. The present data describe the differences in the physical and enzymatic barriers between two species of corynebacteria and also provide some insight into the successful foreign gene expression in corynebacteria.

• Applied Biological Chemistry
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• 제34권2호
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• pp.174-179
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• 1991

Brevibacterium flavum 과 Corynebacterium glutamicum 균주간의 원형질체 융합을 실시하였다. 원항질체 융합을 위하여, Brevibacterium flavum ATCC 21493과 Corynebacterium glutamicum ATCC 21831로부터 여러 변이주를 분리하고 융합의 최적조건을 검토하였다. 본 연구에서 저자 등은 Brevibacterium flavum 108-125와 Corynebacterium glutamicum 41-214A 균주 간의 이속간 원형질체 융합주 MWF 9031의 L-arginine발효능이 우수함을 확인하였다. 융합주 MWF 9031은 10% 포도당이 함유된 배지에서 32.5 mg/ml의 L-arginine을 생산하였다. 융합주의 생리적 성질은 두 모균주의 중간정도를 나타내고, 안정성이 60일 이상 유지되고 있음을 확인하였다.

• Journal of Applied Biological Chemistry
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• 제48권4호
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• pp.218-221
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• 2005

Nucleotide sequence of Brevibacterium flavum ptsG gene capable of complementing Escherichia coli ZSC113 mutations defective to glucose permease activity of phosphotransferase system was completely determined, and the gene product was compared with other glucose-specific enzyme II ($EII^{Glc}$). A ptsG gene of B. flavum consisted of open reading frame of 2,025 nucleotides putatively encoding polypeptide of 675 amino acid residues and TAA stop codon. Deduced amino acid sequence of B. flavum ($EII^{Glc}$) had high homology with ($EIIs^{Glc}$) of Corynebacterium glutamicum, C. efficiens, and B. lactofermentum. Arrangement of structural domains, IIBCA, of B. flanum ($EII^{Glc}$) protein was identical to that of EIIs belonging to glucose-phosphotransferase system.

• 미생물학회지
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• 제23권3호
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• pp.190-196
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• 1985

For frequency improvement of protoplast fusion in Brevibacterium flavum, Brevibacterium lactofermentum lactofermentum and Corynebacterium glutamicum, the effect of plasma expanders on fusion and cell wall regeneration, compatison between direct and two-step selection method, tendency of fusion frequency according to pH of fusion fluid and polyethylene glycol concentration were examined. By addition of 3% polyvinyl pyrrolidone to cell wall regeneration medium, regeneration frequencies were expressed 23 (Brevibacterium lactofermentum), 10.4 (Brevibacterium flavum) and 2.7 (Corynebacterium glutamicum) times higher than those of none polyvinyl pyrrolidone medium respectively.

• 미생물학회지
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• 제26권2호
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• pp.93-100
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• 1988

In order to understand the regulation of glutamate dehydrogenase(GDH) synthesis in Brevibacterium flavum, we have isolated a mutant lacking NADP-linked GDH activity by ethlmethane sulfonate treatment. The $gdh^-$ mutant was grown on the minimal plate with 1mM ammonium chloride and not that with 300mM ammonium chloride. The cell-free extracts from $gdh^-$ mutant and prototroph were also examined with glutamine synthetase(GS) and glutamate synthase (GOGAT) production by niteogen sources. The growth of $gdh^-$ mutant in presence of 20mM ammonium chloride means that GOGAT synthesis is sufficient to allow growth in this condition. GS production of $gdh^-$ mutant as well as parental strain was induced by 1mM urea and ammonium tartrate, but it was repressed by higher concentration of ammonia, and also induced by 20mM to 50mM glutamate as a substrate. It was special attention that GOGAT synthesis from $gdh^-$ strain was more repressed by higher concentration of ammonia than prototroph as described in E. coli system.

• 한국미생물·생명공학회지
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• 제2권2호
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• pp.103-109
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• 1974

전국 각지대의 279개소의 토양시료로 부터 초산자화성세균 383균주를 분리하였으며 이들 초산자화성세균중 L-GA생성 능이 우수한 5균주를 선정하고 균학적성질을 조사하여 Brebacterium flavum nov. sp. D1005B, Corynebacterium glutamicum nov. sp. D1025A, Brevibacterium. flavum nov. sp. D2209B, Corynebacterium acetoacidophilum nov. sp. D2212B 및 Corynebacterium acetoacidophilum nov. sp. D2349A로 동정 명명하였다

• Journal of Microbiology and Biotechnology
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• 제5권4호
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• pp.188-193
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• 1995

A Brevibacterium flavum gene coding for glucose permease of the phosphoenolpyruvate-dependent phosphotransferase system (PTS) was cloned by complementing the Escherichia coli ZSCl13 mutations affecting a ptsG gene with the B. flavum genomic library. From the E. coli clone grown as red colony on a MacConkey plate supplemented with glucose as an additional carbon source, a recombinant plasmid was isolated and named pBFT93. The plasmid pBFT93 was identified as carrying a 3.6-kb fragment of B. flavum chromosomal DNA which enables the E. coli transformant to use glucose or man nose as a sole carbon source in an M9 minimal medium. The non-metabolizable sugar analogues, 2-deoxy-D-glucose (2-DG) and methyl-$\alpha$-D-glucopyranoside (MeGlc) affected the growth of ZSCl13 cells carrying the plasmid pBFT93 on minimal medium supplemented with non-PTS carbohydrate, glycerol, as a sole cabon source, while the analogues did not repress the growth of ZSCl13 cells without pBFT93. It was also found that both $2-deoxy-D-[U-^{14}C]glucose{\;}and{\;}methyl-{\alpha}-D-[U-^{14}C]glucopyranoside$ could be effectively transported into ZSCl13 cells transformed with plasmid pBFT93. Several in vivo complementation studies suggested that the B. flavum DNA in pBFT93 encodes a glucose permease specific for glucose and mannose.

• 한국식품과학회지
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• 제23권5호
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• pp.561-567
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• 1991

본 연구는 Brevibacterium flavum ATCC 14067 및 Corynebacterium glutamicum ATCC 13032간의 protoplast fusion을 행하여 L-methionine의 생산성을 검토하고 발효조건을 개선하기 위하여 연속배양을 행하였다. N-methyl-N'-nitro-N-nitrosoguanidine(MNNG) $500{\mu}g/ml$로 처리하였으며 B. flavum K-104($Thr\;Met\;Km^{r}\;Et^{r}\;Sm^{r}\;Tm^{r}$)와 C. glutamicum B-70($Thr\;Hos\;Km^{r}\;Et^{r}\;Sm^{r}\;Tm^{r}$)의 변이주를 분리하였다. 이들 변이주에 $500{\mu}g/ml$의 lysozyme을 처리하였을 때 원형질체 형성을 및 재생율은 각각 99% 및 $64{\sim}66%$를 나타내었으며 융합 빈도는 3% PVP를 함유한 35% PEG 용액에서 $3.5{\times}10^{5}$을 나타내었다. Sodium alginate로 고정화시킨 융합주 BFCG 37은 72시간 회분배양에서 0.89g/l의 methionine을 생산하였고 연속배양에서는 $18.75mg/^{1}hr\;^{1}$의 L-methionine를 안정적으로 생산할 수 있었다.