• Journal of Microbiology and Biotechnology
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• 제7권5호
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• pp.360-362
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• 1997

A transposon Tn5 mutant of Rhodobacter sphaeroides 2.4.1 was isolated for its impaired ability of growth on minimal medium containing ${\beta}$-hydroxybutyric acid as a sole carbon source. The mutant, R. sphaeroides S7 showed approximately 6-fold decrease in ${\beta}$-hydroxybutyrate dehydrogenase activity compared with that of wild type. In R. sphaeroides S7 the Tn5 was located in DNA region corresponding to a 4.2-kb EcoRI DNA fragment of R. sphaeroides 2.4.1 chromosome.

• Journal of Applied Biological Chemistry
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• 제48권4호
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• pp.183-186
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• 2005

Arabidopsis AHL gene encodes a 3'(2')-phosphoadenosine 5'-phosphate (PAP)-specific phosphatase that plays a role in the sulfate activation pathway. We complemented E. coli cysQ mutant defective in cysteine biosynthesis with the AHL gene. AHL cDNA was cloned into the prokaryotic expression vector pKK388-1 and transformed into the bacterial mutant. Since cysQ mutant is a leaky cysteine auxotroph only under aerobic conditions, the bacteria were grown in liquid media with vigorous shaking to provide more aeration. In cysteine-free medium, cysQ mutant and the mutant harboring empty vector did not grow well, whereas cells harboring AHL cDNA exhibited significantly improved growth with doubling time of approximately 3 h. cysQ is known to encode a 3'(2'),5'-diphosphonucleoside 3'(2')-phosphohydrolase (DPNPase). However, our data suggest that cysQ protein has PAP-specific phosphatase activity in addition to DPNPase activity. Microbial complementation procedure described in this paper is useful for structure-activity studies of PAP-specific phosphatases identified from microbes and plants.

• BMB Reports
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• 제30권3호
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• pp.167-172
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• 1997

The yeast Saccharomyces cerevisiae, mutant CH117, shows a drug-hypersensitivity (dhs) to cycloheximide, bleomycin, actinomycin D, 5-fluorouracil. nystatin, nigericin and several other antibiotics. CH 117 was also temperature-sensitive (ts). being unable to grow at $37^{\circ}C$ and secreted more invertase and acid phosphatase into the medium than the parent yeast. CH117 grows very slowly and the cell shape is somewhat larger and more sensitive to zymolyase than the wild type cells. Light microscopic and electron microscopic observation also revealed abnormality of the mutant cell wall. These characteristics indicate that CH117 has a defect in an essential component of the cell surface and that the cell wall which performs barrier functions has become leaky in the mutant. We screened a genomic library of wild type yeast for clones that can complement the mutation of CH117. A plasmid, pCHX1, with an insert of 3.6 kilobases (kbs) could complement the dhs and ts of CH117. Deletion and subcloning of the 3.6 kb insert showed that a gene for the complementation of mutant phenotypes was located in 1.9 kbs Puvll-Hindlll fragment.

• 방사선산업학회지
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• 제4권1호
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• pp.65-71
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• 2010

Two chitinase producing strains CHI2 and CHI4 were isolated from soybean rhizosphere soil. Both the strains belonged to Lysobacter enzymogenes as indicated by 16S rDNA sequence analysis. Though strain CHI2 and CHI4 produced extracellular chitinase, they differ in their chitinolytic activity. CHI4 produced approximately three times the higher amounts of enzyme than that of CHI2 under specified conditions. CHI2 produced $535.67U\;l^{-1}$ of chitinase after 48 h incubation with a specific activity of $3.91U\;mg^{-1}$ of protein while strain CHI4 produced $1584.13U\;l^{-1}$ of chitinase with a specific activity of $10.88U\;mg^{-1}$ protein. SDS-PAGE analysis indicated that the molecular weight of chitinase enzyme was approximately 45 kDa. A faint band with a molecular weight of 55 kDa reveals the possibility for the presence of another kind of chitin binding protein. Mutant library was developed by exposing the isolates to gamma rays at their $LD_{99}$ value (0.23 kGy). Totally, 11 mutants of CHI2 and CHI4 are reported to have enhanced chitinase activity. Several leaky mutant clones with decreased enzyme activity and a defective mutant (CHI2-M16) with complete loss of chitinase activity were also identified. CHI4-M18, CHI4-M8 and CHI4-M29 showed 78.8, 41.5, and 31.9% increased chitinase activity over wild type CHI4.

• Journal of Microbiology and Biotechnology
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• 제10권2호
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• pp.169-175
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• 2000

The filamentous soil bacterium Streptomyces coelicolor is known to produce four distinct antibiotics. The simultaneous global regulation for the biosynthesis of those four antibiotics was previously confirmed by absA and absB mutations that blocked all four antibiotics' biosynthesis without influencing their morphological differentiation. To study the complex regulatory cascade that controls the secondary metabolism in Streptomyces, a new abs-like mutation was characterized. namely absR, which is slightly leaky on a complete R2YE medium, yet tight on a minimal medium. A genetic analysis of the absR locus indicated that it is located at 10 o'clock on the genetic map, near the site of absA. A cloned copy of the absA gene that encoded bacterial two-component regulatory kinases did not restore antibiotic biosyntheis to the absR mutant. Accordingly, it is proposed that absR is another abs-type mutation which is less tight than the previously identified absA or absB mutations income medium conditions, and can be used to characterize another global regulatory gene for secondary metabolete formation in S. coelicolor.

• 미생물학회지
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• 제44권3호
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• pp.193-198
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• 2008

ErmSF는, 235 rRNA의 A2058에 methylation 시켜서 macrolide-lincosamide-streptogramin B ($MLS_B$)계 항생제의 부착을 저해함으로써 항생제의 활성을 억제하는 내성인자 단백질인 ERM 단백질의 하나로, 다른 ERM 단백질과는 달리 긴 N-terminal end region을 가지고 있고 이 부위의 25%를 arginine이 차지하고 있다. 특히 $^{58}RARR^{61}$ 부위에 arginine이 모여 있어서 여기에 존재하는 R의 역할을 알아보기 위해 1-57, 1-59 그리고 1-60과 1-61이 제거된 결손 변이 단백질을 대장균에 발현하고 그 활성을 성장곡선을 작성하여 알아보았다. 그 결과 R60과 R6l이 활성에 중요한 것으로 관찰되었다. 1-59의 아미노산이 결손 된 유전자를 사용하여 R60A, R61A와RR60, 61AA의 위치선정 치환 변이 단배질의 세포내 활성을 측정하여 본 결과 R60의 역할이 R6l보다 큰 것으로 관찰되었으며 이들의 활성에서의 역할은 상호보완적인 것으로 나타났다. 그리고 이 아미노산들이 2차 구조인 $\alpha$-helix의 일부일 것으로 추정되었다.

• The Plant Pathology Journal
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• 제33권1호
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• pp.53-65
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• 2017

Barley yellow dwarf virus (BYDV) belongs to Luteovirus and is limited only at phloem related tissues. An open reading frame (ORF) 4 of BYDV codes for the movement protein (MP) of BYDV gating plasmodesmata (PD) to facilitate virus movement. Like other Luteoviruses, ORF 4 of BYDV is embedded in the ORF3 but expressed from the different reading frame in leaky scanning manner. Although MP is a very important protein for systemic infection of BYDV, there was a little information. In this study, MP was characterized in terms of subcellular localization and programmed cell death (PCD). Gene of MP or its mutant (ΔMP) was expressed by Agroinfiltration method. MP was clearly localized at the nucleus and the PD, but ΔMP which was deleted distal N-terminus of MP showed no localization to PD exhibited the different target with original MP. In addition to PD localization, MP appeared associated with small granules in cytoplasm whereas ΔMP did not. MP associated with PD and small granules induced PCD, but ΔMP showed no association with PD and small granules did not exhibit PCD. Based on this study, the distal N-terminal region within MP is seemingly responsible for the localization of PD and the induction small granules and PCD induction. These results suggest that subcellular localization of BYDV MP may modulate the PCD in Nicotiana benthamiana.