• Proceedings of the Korean Society for Bioinformatics Conference
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• 2003.10a
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• pp.277-287
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• 2003

Acetolactate synthase (ALS, EC 4.1.3.18; also referred to as acetohydroxy acid synthase) catalyzes the first common step in the biosynthesis of valine, leucine, and isoleucine in microorganisms and plants. Recently X-ray structure of yeast ALS was available. Pair-wise alignment of yeast and tobacco ALS sequences revealed 63% sequence similarity. Using Deep View and automatic modeling on Swiss model server, we have generated reliable models of tobacco ALS based on yeast ALS template with a calculated pair-wise RMSD of 0.86 Angstrom. Functional roles of four residues located on the subunit interface (H142, El43, M350, and R376) were examined by site-directed mutagenesis. Seven mutants were generated and purified, of which three mutants (H142T, M350V, and R376F) were found to be inactivated under various assay conditions. The H142k mutant showed moderately altered kinetic properties. The E143A mutant increased 10-fold in K$_m$ value while other parameters remained unchanged. The M350C mutant was strongly resistant to three tested herbicides, while the R376k mutant can bind with herbicide carder at similar affinity to that of wild type enzyme, as determined by tryptophan quenching study. Except M350V mutant, all other mutants were ate to bind with cofactor FAD. Taken together, it is likely that residues H142 and E143 are located at the active site, while residues M350 and R376 are possibly located at the overlapping region of active site and herbicide binding site of the enzyme. Our data also allows us to hypothesize that the interaction between side chains of residues M350 and R376 are probably essential for the correct conformation of the active site. It remains to be elucidated that, whether the herbicide, upon binding with enzyme, inactivates the enzyme by causing change in the active site allosterically, which is unfavorable for catalytic activity.

• Journal of Microbiology and Biotechnology
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• v.28 no.9
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• pp.1573-1579
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• 2018

Transcriptional gene silencing is regulated by the chromatin structure, which is by various factors including histones. Saccharomyces cerevisiae contains transcriptionally silenced regions such as telomeric regions and hidden mating (HM) loci. The positively-charged amino acids on the histone H4 tail were reported to be critical for the telomeric silencing in yeast, by interacting with Dot1, a specific methyltransferase for the $79^{th}$ lysine on histone H3. However, Dot1 did not affect gene silencing within HM loci, but whether the positively-charged amino acids on the H4 tail affect HM silencing has not been defined. To elucidate the function of the H4 tail on HM silencing, we created several MATa-type yeast strains bearing the substitution of arginine with alanine or lysine on the histone H4 tail and checked the sensitivity of MATa-type yeast to alpha pheromone. The arginine point mutants substituted by alanine (R17A, R19A, and R23A) did not show sensitivity to alpha pheromone, but only two arginine mutants substituted by lysine (R17K and R19K) restored the sensitivity to alpha pheromone-like wild type. These data suggested that the basic property of arginine at $17^{th}$ and $19^{th}$ positions in the histone H4 tail is critical for maintaining HM silencing, but that of the $23^{rd}$ arginine is not. Our data implicated that the positive charge of two arginine residues on the histone H4 tail is required for HM silencing in a manner independent of Dot1.

• Applied Biological Chemistry
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• v.31 no.1
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• pp.26-32
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• 1988

This study was performed to construct killer wine yeasts which might suppress the growth of wild yeasts, reduce the consumption of starter and condense the fermentation period. Saccharomyces cerevisiae M524, a commercial wine yeast, was treated with N-methyl-N'-nitro-N-nitrosoguanidine to induce auxotrophic mutants, i.e., CHM $2(thr^-)$, CHM 3 $(asp^-)$ and CHM 6 $(tyr^-)$. These auxotrophs were fused successfully with a killer yeast, S. cerevisiae $1368R({\alpha}\;his\;4\;kar\;1-1(kil-k)\;(k_0)$, respiratory deficient) using sphoroplast techniques and the fusants were designated as CHF 21$(th^-\;kil^+)$, CHF 22$(thr^-\;kil^+)$, CHF 31$(asp^-\;kil^+)$ and GHF 61$(tyr^-\;kil^+)$. Combined cultivation of CHF 31 with 1368R or S. cerevisiae $5{\times}47$ (killer sensitive) proved out that CHF 31 had the characteristic of killing and produced the same amount of ethanol as the prototroph, M524.

• Korean Journal of Environmental Biology
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• v.34 no.3
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• pp.177-182
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• 2016

Malodor emitted while producing fertilizer from hatchery egg waste treated with microorganism is an important limiting factor. To reduce this problem, we attempted to use two yeast strains, Saccharomyces cerevisiae, KACC 30008 and KACC 30068. Both yeast strains reduce ammonia gas emission 35.4% than only treated with bacterium, Bacillus amyloliquefaciens. When both strains were used together, that was reduced as 57.1%. KACC 30008 and 30068 strains reduced hydrogen sulfide 42 and 90.4%, respectively. Both strains together reduced hydrogen sulfide gas as 98.5%. KACC 30008 did not decrease methyl mercaptan emission. However KACC 30068 decreased 40% and both strains together decreased the gas emission as 66.7%. Overall, this study showed that yeast treatment could enhance the effect of B. amyloliquefaciens treatment in the reduction of malodorous gas emission.

• Molecules and Cells
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• v.22 no.2
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• pp.146-153
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• 2006

Cell polarity is critical for the division, differentiation, migration, and signaling of eukaryotic cells. RAX2 of budding yeast encodes a membrane protein localized at the cell cortex that helps maintain the polarity of the bipolar pattern. Here, we designate SPAC6f6.06c as $rax2^+$ of Schizosaccharomyces pombe, based on its sequence homology with RAX2, and examine its function in cell polarity. S. pombe $rax2^+$ is not essential, but ${\Delta}rax2$ cells are slightly smaller and grow slower than wild type cells. During vegetative growth or arrest at G1 by mutation of cdc10, deletion of $rax2^+$ increases the number of cells failing old end growth just after division. In addition, this failure of old end growth is dramatically increased in ${\Delta}tea1{\Delta}rax2$, pointing to genetic interaction of $rax2^+$ with $tea1^+$. ${\Delta}rax2$ cells contain normal actin and microtubule cytoskeletons, but lack actin cables, and the polarity factor for3p is not properly localized at the growing tip. In ${\Delta}rax2$ cells, and endogenous rax2p is localized at the cell cortex of growing cell tips in an actin- and microtubule-dependent manner. However, ${\Delta}rax2$ cells show no defects in cell polarity during shmoo formation and conjugation. Taken together, these observations suggest that rax2p controls the cell polarity of fission yeast during vegetative growth by regulating for3p localization.

• The Korean Journal of Mycology
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• v.51 no.4
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• pp.349-359
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• 2023

Freshwater ecosystems are significant habitats of fungi including yeasts. This study aimed to isolate and characterize wild yeasts from freshwater environments in Korea. The yeast isolates were identified by using the D1/D2 domains of the 26S rDNA regions. We identified four strains, Candida viswanathii (NNIBRFG39781), Curvibasidium cygneicollum (NNIBRFG49003), Oberwinklerozyma silvestris (NNIBRFG39803) and Vishniacozyma foliicola (NNIBRFG6120). These yeasts had not previously been recorded in Korea. We investigated the morphological and cultural characteristics of these yeasts. All of them grew on YPD, PD and YM media. Candida viswanathii (NNIBRFG39781), O. silvestris (NNIBRFG39803) and V. foliicola (NNIBRFG6120) grew in YPD medium containing glucose and in pH range of 4-8. Curvibasidium cygneicollum (NNIBRFG49003) grew well low temperature compared to others and slowly.

• The Korean Journal of Mycology
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• v.44 no.1
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• pp.1-7
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• 2016

This study focused on isolation and identification of wild yeasts from soils in fields near mountains and elucidation of its yeast distribution. Several kinds of yeasts were isolated from various soils of Daejeon metropolitan city and Chungcheongnam-do in Korea and identified by BLAST search of nucleotide sequences of internal transcribed spacer (ITS) region including 5.8S rRNA and D1/D2 region of 26S rDNA. Ninety-seven strains of 20 species from 61 soil samples were isolated, of which Cryptococcus podzolicus (11 strains), Debaryomyces hansenii (6 strains), and Trichosporon asahii (6 strains) were dominant species.

• The Plant Pathology Journal
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• v.25 no.2
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• pp.193-198
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• 2009

A full-length cDNA of OgGRP gene encoding a glycinerich cell wall protein was isolated from wild rice (Oryza grandiglumis). Deduced amino acid sequences of OgGRP are composed of 148 amino acids (16.3 kDa), and show 85.9% homology with Osgrp-2 (Oryza sativa). RT-PCR analysis showed that RNA expression of OgGRP was regulated by defense-related signaling chemicals, such as cantharidin, endothall, jasmonic acid, wounding, or yeast extract treatment. In relation to pathogen stress, the function of OgGRP was analyzed in OgGRP over-expressing Arabidopsis thaliana. Overexpression of OgGRP in Arabidopsis contributed to moderate resistance against fungal pathogen, Botrytis cinerea, by lowering disease rate and necrosis size. In the analysis of the transgenic Arabidopsis lines to check the change of gene expression profile, induction of PR1, PR5 and PDF1.2 was confirmed. The induction seemed to be caused by the interaction of ectopic expression of OgGRP with SA-and JA-dependent signaling pathways.

• The Plant Pathology Journal
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• v.19 no.1
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• pp.24-29
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• 2003

SS2-2, a hypernodulating soybean mutant was isolated by EMS mutagenesis from Sinpaldalkong 2. This auto-regulation mutant showed greater number of nodules and smaller plant size than its wild type Sinpaldalkong 2. SSR markers were used to identify DNA variation at SSR loci from different soybean LG. The only SSR marker that detected a length polymorphism between SS2-2 and its wild type ancestor was Satt294 on LG C1 instead of LG H, locating a hypernodulating gene. Sequencing data of flanking Satt294 indicated that the size variation was due to extra stretch of TTA repeats of the SSR motif in SS2-2, along with $A\longrightarrow$G transversion. In spite of phenotypic differences between the wild type and its hypernodulating mutants, genomic DNA poly-morphisms at microsatellite loci could not control regulation of nodule formation. The cDNA-AFLP method was applied to compare differential display of cDNA between Sinpaldalkong 2 and SS2-2. After isolation and sequence comparison with many AELP fragments, several interesting genes were identified. Northern blot analysis, immunolocalization and/or the yeast two-hybrid system with these genes might provide information on regulation of nodule development in SS2-2.

• Bulletin of the Korean Chemical Society
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• v.22 no.5
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• pp.514-518
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• 2001

We have explored the importance of two ATP binding domains of Hsp104 protein in protection of yeast cells from cadmium exposure. In the previous study we have discovered that the presence of two ATP binding sites was essential in providing heat sh ock protection as well as rescuing cells from oxidative stress. In this paper we first report wild type cell with functional hsp104 gene is more resistant to cadmium stress than hsp104-deleted mutant cell, judging from decrease in survival rates as a result of cadmium exposure. In order to demonstrate functional role of two ATP binding sites in cadmium defense, we have transformed both wild type (SP1) and hyperactivated ras mutant (IR2.5) strains with several plasmids differing in the presence of ATP binding sites. When an extra copy of functional hsp104 gene with both ATP binding sites was overexpressed with GPD-promoter, cells showed increased survival rate against cadmium stress than mutants with ATP binding sites changed. The degree of protection in the presence of two ATP binding sites was similarly observed in ira2-deleted hyperactivated ras mutant, which was more sensitive to oxidative stress than wild type cell. We have concluded that the greater sensitivity to cadmium stress in the absence of two ATP binding sites is attributed to the higher concentration of reactive oxygen species (ROS) produced by cadmium exposure based on the fluorescence tests. These findings, taken all together, imply that the mechanism by which cadmium put forth toxic effects may be closely associated with the oxidative stress, which is regulated independently of the Ras-cAMP pathway. Our study provides a better understanding of cadmium defense itself and cross-talks between oxidative stress and metal stress, which can be applied to control human diseases due to similar toxic environments.