• Bulletin of the Korean Chemical Society
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• 제34권6호
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• pp.1673-1678
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• 2013

Lumazine protein is a member of the riboflavin synthase superfamily and the intense fluorescence is caused by non-covalently bound to 6,7-dimethyl 8-ribityllumazine. To figure out the binding modes and the structure of the N-terminal domain of lumazine protein, the wild type of protein extending to amino acid 118 (N-LumP 118 Wt) and mutants of N-LumP 118 V41W, S48W, T50W, D64W, and A66W from Photobacterium leiognathi were purified. The biochemical properties of the wild type and mutants of N-LumP 118 proteins were analyzed by absorbance and fluorescence spectroscope. The peak of absorbance and fluorescence of lumazine ligand were shifted to longer wavelength on binding to N-LumPs. The observed absorbance value at 410 nm of lumazine bound to N-LumP 118 proteins indicate that one mole of N-LumP 118 proteins bind to one mole of ligand of lumazine. Fluorescence analysis show that the maximum peak of fluorescence of N-LumP S48W was shifted to the longest wavelength by binding with 6,7-dimethyl 8-ribityllumazine and was shown to the greatest quench effect by acrylamide among all tryptophan mutants.

• Journal of Microbiology
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• 제42권3호
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• pp.194-199
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• 2004

Investigation of the expression of the riboflavin (rib) genes, which are found immediately downstream of luxG in the lux operon in Photobacterium phosphoreum, provides more information relevant to the evolution of bioluminescence, as well as to the regulation of supply of flavin substrate for bacterial bioluminescence reactions. In order to answer the question of whether or not the transcriptions of lux and rib genes are integrated, a transcriptional termination assay was performed with P. phoxphoreum DNA, containing the possible stem-loop structures, located in the intergenic region of luxF and luxE ($\Omega$$\_$A/), of luxG and ribE ($\Omega$$\_$B/), and downstream of ribA ($\Omega$$\_$c/). The expression of the CAT (Chloram-phenicol Acetyl Transferase) reporter gene was remarkably decreased upon the insertion of the stem-loop structure ($\Omega$$\_$c/) into the strong lux promoter and the reporter gene. However, the insertion of the structure ($\Omega$$\_$B/) into the intergenic region of the lux and the rib genes caused no significant change in expression from the CAT gene. In addition, the single stranded DNA in the same region was protected by the P. phosphoreum mRNA from the Sl nuclease protection assay. These results suggest that lux genes and rib genes are part of the same operon in P. phosphoreum.

• Fisheries and Aquatic Sciences
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• 제3권1호
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• pp.52-63
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• 2000

Non-specific reaction has been a problem in doing, especially, research and diagnosis for infectious agents. Avidin-biotin-peroxidase complex (ABC) techniques has widely been used to amplify a reaction. Photobacterium damse1a subsp. piscicdia (formerly Pasteurella piscicida) exhibited a capacity to bind with streptavidin non-specifically. The band, estimated 26 K Da in Western blotted paper, was blocked with biotin but incompletely. In an attempt to explore an involvement of the non-specific substance in attaching piscine cells, cell attachment test performed using anti- Ph. d. subsp piscicida sera raised mouse and rabbit exhibited slightly blocking effects for Mediterranean (1736) and significantly for Japanese (Sp 92144) isolate. Biotin decreased the attachment ability significantly for Sp92144 but it was not effective to 1736. Both isolates showed greatly enhanced attachment ability with poly-L-lysin. The non-specific binding substance was contained in bacterial extracellular products (ECPs). The substance was able to purified with 2-imminobiotin affinity column, the purified substance appeared to have 4 bands in silver staining, and had a carbohydrate branch. This purified substance showed cytotoxic effects selectively between 5 piscine cell lines. Moreover, it stimulated rainbow trout macrophage in terms of reduction of cytochrome cas well as yeast phagocytosis, significantly.

• KSBB Journal
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• 제14권4호
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• pp.403-407
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• 1999

식품이나 수질 내의 독성 물질 monitoring을 위해서 발광미생물인 P. phosphoreum이 많이 연구되고 있는데 이 독성 물질측정을 위하여 P. phosphoreum을 더 효과적으로 이요하기 위해 고정화하여 이용하는 방법을 연구하였다. 고정화 방법을 크게 4가지로 나누어서 그 방법에 따라 각각 고정화 물질 한가지씩을 선택하여 P. phosphoreum의 bioluminescence 안정성을 알아보았다. Polvacrylamide나 collagen에서는 bioluminescence가 유지를 못하고 material과 cell을 혼합하자마자 급격히 떨어졌으나 alginate와 k-carrageenan에서는 빛 안정성이 매우 좋았다. 그러나 k-carrageenan은 온도를 높여야 gel이 형성되는 성질을 갖고 있기 때문에 저온성 발광 미생물인 P. phosphoreum에는 적합한 고정화 물질이 되지 못한다. 따라서 P. phosphoreum의 bioluminescence를 안정되게 유지하면서 고정화가 용이한 polymer로는 alginate가 적합하다.

• 한국미생물·생명공학회지
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• 제47권1호
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• pp.139-147
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• 2019

Glutamate decarboxylase catalyzes the conversion of glutamate to gamma-aminobutyric acid (GABA), contributing to pH homeostasis through proton consumption. The reaction is the first step toward the GABA shunt. To date, the enzymes involved in the glutamate metabolism of Photobacterium damselae subsp. piscicida have not been elucidated. In this study, an open reading frame of P. damselae subsp. piscicida, showing homology to the glutamate decarboxylase or putative pyridoxal-dependent aspartate 1-decarboxylase genes, was isolated and cloned into an expression vector to produce the recombinant enzyme. Preliminary gas chromatography-mass spectrometry characterization of the purified recombinant enzyme revealed that it catalyzed not only the decarboxylation of glutamate but also the transamination of GABA. This enzyme of P. damselae subsp. piscicida could be bifunctional, combining decarboxylase and transaminase activities in a single polypeptide chain.

• 한국식품영양과학회지
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• 제45권3호
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• pp.396-401
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• 2016

Histamine을 생성하는 Morganella morganii와 Photobacterium phosphoreum으로부터 crude histidine decarboxylase(HDC)를 추출하여 $65{\sim}121^{\circ}C$로 열처리한 다음 균의 생육 및 효소 활성 변화에 대해 살펴보았다. 그 결과 M. morganii와 P. phosphoreum은 모든 열처리 조건에서 비 가열 처리구와 비교 시 균의 생육이 저해됨을 확인하였다. M. morganii와 P. phosphoreum 유래 HDC의 효소 활성은 $65^{\circ}C$에서 90% 이상의 효소 활성이 저해됨을 확인하였고, 온도가 증가함에 따라 유의적으로 활성이 억제되는 것으로 나타났다. SDS-PAGE 결과에서는 $65{\sim}100^{\circ}C$ 범위에서 비가열 처리구와 비교 시 조효소액의 단백질 패턴의 변화가 크지 않았으나, $121^{\circ}C$에서 단백질 band의 강도가 크게 약해졌다. Native-PAGE에서는 $65^{\circ}C$ 처리 조건에서부터 단백질 패턴의 변화가 크게 나타났다. 따라서 가열처리($65{\sim}121^{\circ}C$)는 histamine 생성균인 M. morganii와 P. phosphoreum의 생육을 억제할 뿐만 아니라 HDC의 효소 활성도가 저해됨을 확인하여, 식품산업에서 적용되고 있는 열처리 조건에서 histamine 생성 억제에 큰 효과가 있는 것으로 생각한다.

• Biotechnology and Bioprocess Engineering:BBE
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• 제8권2호
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• pp.147-150
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• 2003

Water toxicity monitoring based on the continuous cultivation of Photobacterium phorphoreum is presented. Normally, after 10 days of operation, a dark variant, which emits no light, appears and dominates the population, resulting in a rapid decrease in bioluminescence. Therefore, to overcome this problem, a fluidized-bed reactor is used in which alginate-immobilized cells are grown and leaking cells are continuously released into the effluent Experimental results revealed that the dominance of dark variants was suppressed inside the immobilized beads, thereby mitigating the rapid loss of bioluminescence. Plus, a high dilution rate (1.2 h$\^$-1/) prevented the occurrence of other microbial contamination in the reactor The concentration and bioluminescence of the released cells were sufficient to measure the water toxicity for more than 4 weeks.

• BMB Reports
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• 제33권4호
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• pp.353-357
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• 2000

A new assay method of ${\alpha}-Oxidase$ (fatty acid : oxygen dioxygenase, 1-decarboxylating) was developed using a bioluminescence reaction system of marine luminous bacterium, Photobacterium phosphoreum. ${\alpha}$-Oxidase was isolated from a cucumber (Cucumis sativus). Pentadecanoic acid was used as a substrate, and the product, tetradecanal, was analyzed with a bacterial luciferase-coupled reaction. Initial light intensity was directly related to the concentration of tetradecanal in the range of 1 nM to 10 ${\mu}M$. Optimal pH and temperature were 7.5 and $25^{\circ}C$, respectively. Optimal pentadecanoic acid concentration in a standard assay of ${\alpha}$-oxidase was 0.1 mM. The Km value of pentedecanoic acid was $85{\mu}M$. This method is straightforward, rapid, convenient, and easy. Its needs no treatment or extraction of reaction mixture.

• Journal of Microbiology and Biotechnology
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• 제7권4호
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• pp.242-249
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• 1997

Stability of bioluminescence was investigated with Photobacterium phosphoreum immobilized on the strontium alginate in order to develope continuous real time monitoring of pollutants. The stability of bioluminescence emission was improved by prolonged aging time. The aging time of ${\geq}40$ min and the cell concentration of ${\leq}0.6\;of\;OD_660$ were selected for the immobilization of P. phosphoreum to give linearity between cell concentrations and bioluminescence intensity. In sensitivity tests using phenol, it was found that this compound quenched bioluminescence proportional to the concentration without lowering of cell growth. The lower value for maximum quenching ($q_s$) and higher dissociation constant ($K_s$) were observed with strontium-alginate immobilized cells compared to free cells. The response of bioluminescence to toxicants was evaluated with the immobilized luminescent bacteria. The sensitivity of the immobilized cells was found to be good in response to toxicants, 4-nitrophenol, salicylate and cadmium, when evaluated with a specific rate of bioluminescence quenching.

• 환경생물
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• 제18권4호
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• pp.441-446
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• 2000

Trichoderma harzianum is an aggressive causal agent of green mold disease on mushroom cultivation. Some bacterial strains isolated, from oyster mushroom compost in Wonju, were found to have in vitro antifungal activity against Trichoderma harzianum ATCC 6385, 6504, and our isolates Trichoderma spp. Y and G. Further in vitro antifungal studies on several strains of phytopathogenic fungi showed that all of 12 phytopathogenic fungal strains were significantly inhibited by the isolated antifungal bacteria in Petri dishes. Of these, KATB 99121 showed the broadest inhibiting effect and displayed as negative coagulase, negative sulfide production and rod shape. KATB 99121 was resistant to ampicillin, chlorampenicol, and kanamycin. Identification of isolates was determined by Biolog GN system, and KATB 99121 was identified as Photobacterium logei because of 96 probability, 0.65 similarity, and 4.97 disturbance. With electron microscopy, thin section of KATB 99121 strain revealed typical rod-like shaped cell (0.6-0.8${\mu}{\textrm}{m}$$\times$1.5-2.0${\mu}{\textrm}{m}$) with prokaryotic structure and organization.