• Mycobiology
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• v.41 no.2
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• pp.67-72
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• 2013

Pathogenic microbes secrete various enzymes with lipolytic activities to facilitate their survival within the host. Lipolytic enzymes include extracellular lipases and phospholipases, and several lines of evidence have suggested that these enzymes contribute to the virulence of pathogenic fungi. Candida albicans and Cryptococcus neoformans are the most commonly isolated human fungal pathogens, and several biochemical and molecular approaches have identified their extracellular lipolytic enzymes. The role of lipases and phospholipases in the virulence of C. albicans has been extensively studied, and these enzymes have been shown to contribute to C. albicans morphological transition, colonization, cytotoxicity, and penetration to the host. While not much is known about the lipases in C. neoformans, the roles of phospholipases in the dissemination of fungal cells in the host and in signaling pathways have been described. Lipolytic enzymes may also influence the survival of the lipophilic cutaneous pathogenic yeast Malassezia species within the host, and an unusually high number of lipase-coding genes may complement the lipid dependency of this fungus. This review briefly describes the current understanding of the lipolytic enzymes in major human fungal pathogens, namely C. albicans, C. neoformans, and Malassezia spp.

• Journal of Microbiology and Biotechnology
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• v.17 no.10
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• pp.1655-1660
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• 2007

A metagenome is a unique resource to search for novel microbial enzymes from the unculturable microorganisms in soil. A forest soil metagenomic library using a fosmid and soil microbial DNA from Gwangneung forest, Korea, was constructed in Escherichia coli and screened to select lipolytic genes. A total of seven unique lipolytic clones were selected by screening of the 31,000-member forest soil metagenome library based on tributyrin hydrolysis. The ORFs for lipolytic activity were subcloned in a high copy number plasmid by screening the secondary shortgun libraries from the seven clones. Since the lipolytic enzymes were well secreted in E. coli into the culture broth, the lipolytic activity of the subclones was confirmed by the hydrolysis of p-nitrophenyl butyrate using culture supernatant. Deduced amino acid sequence analysis of the identified ORFs for lipolytic activity revealed that 4 genes encode hormone-sensitive lipase (HSL) in lipase family IV. Phylogenetic analysis indicated that 4 proteins were clustered with HSL in the database and other metagenomic HSLs. The other 2 genes and 1 gene encode non-heme peroxidase-like enzymes of lipase family V and a GDSL family esterase/lipase in family II, respectively. The gene for the GDSL enzyme is the first description of the enzyme from metagenomic screening.

• Proceedings of the Korean Society of Dyers and Finishers Conference
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• 2008.10a
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• pp.29-30
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• 2008

• Korean Journal of Microbiology
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• v.30 no.3
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• pp.171-186
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• 1992

Lipolytic characteristics of candia cylindracea lipase was studied by various triacylglycerols with different fatty acyl chains as substrate. The substrate was emulsified with gum arabic and the rate of hydrolysis was determined by pH stat method. The effects of gum concentration, pH, temperature, and $Ca^{2+}$ ion on the enzyme activities were examined. The results show that the effect of these factors are markedly depending on the structurla nature of substrates. The triolein was the best substrate among tested. Present study demonstrates that for characterization of lipolytic enzymes, it is critically important to select proper substrate and activator.r.

• Journal of Microbiology and Biotechnology
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• v.14 no.5
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• pp.952-958
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• 2004

Of 23 psychrotrophic bacteria isolated from the west Pacific deep-sea sediments, 19 were assigned to the $\gamma$-Proteobacteria, 3 to the <$\beta$-Proteobacteria, and 1 to the Gram-positive bacteria, as determined by their 16S rDNA sequences. Ten psychrotrophs, affiliated to the Psychrobacter, Pseudoalteromonas, and Pseudomonas genera in the $\gamma$-Proteobacteria group, were screened for lipolytic bacteria. The majority of the lipolytic isolates had growth temperatures between 4-$30^\circ{C}$, and all of them were neutrophilic, aerobic, or facultatively anaerobic, and some were able to produce multiple kinds of ectohydrolytic enzymes. The deep-sea strains Psychrobacter sp. wp37 and Pseudoalteromonas sp. wp27 were chosen for further lipase production analysis. Both strains had the highest lipase production when grown at 10 to $20^\circ{C}$; their highest lipase production occurred at the late-exponential growth stage; and the majority of the enzymes were excreted to the outside of the cells. Lipases from both strains had the same optimal reaction temperature and pH (20-$30^\circ{C}$, pH 7-8) and could retain about 60% of their highest activity at $4^\circ{C}$. Furthermore, SDS-PAGE and an in-gel activity test showed that they had the same high molecular mass of about 85 kDa.

• Journal of Microbiology and Biotechnology
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• v.24 no.6
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• pp.771-780
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• 2014

A putative lipolytic enzyme gene, named as est9x, was obtained from a marine microbial metagenome of the South China Sea. Sequence analysis showed that Est9X shares lower than 27% sequence identities with the characterized lipolytic enzymes, but possesses a catalytic triad highly conserved in lipolytic enzymes of the ${\alpha}/{\beta}$ hydrolase superfamily. By phylogenetic tree construction, Est9X was grouped into a new lipase/esterase family. To understand Est9X protein in depth, it was recombinantly expressed, purified, and biochemically characterized. Within potential hydrolytic activities, only lipase/esterase activity was detected for Est9X, confirming its identity as a lipolytic enzyme. When using p-nitrophenol esters with varying lengths of fatty acid as substrates, Est9X exhibited the highest activity to the C2 substrate, indicating it is an esterase. The optimal activity of Est9X occurred at a temperature of $65^{\cric}C$, and Est9X was pretty stable below the optimum temperature. Distinguished from other salt-tolerant esterases, Est9X's activity was tolerant to and even promoted by as high as 4 M NaCl. Our results imply that Est9X is a unique esterase and could be a potential candidate for industrial application under extreme conditions.

• Proceedings of the Korean Society of Dyers and Finishers Conference
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• 2008.04a
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• pp.210-212
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• 2008

• Proceedings of the Korean Society of Dyers and Finishers Conference
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• 2010.03a
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• pp.163-164
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• 2010

• BMB Reports
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• v.45 no.1
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• pp.14-19
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• 2012

A feruloyl esterase encoding gene (designated fae6), derived from a leachate metagenomic library, was cloned and the nucleotide sequence of the insert DNA determined. Translational analysis revealed that fae6 consists of a 515 amino acid poly-peptide, encoding a 55 kDa pre-protein. The Fae6 primary structure contained the G-E-S-A-G sequence, which corresponds well with a typical catalytic serine sequence motif (G-x-S-x-G). The fae6 gene was successfully over-expressed in E. coli and the recombinant protein was purified to 8.4 fold enrichment with 17% recovery. The $K_M$ data showed Fae6 has a high affinity to methyl sinapate while thermostability data indicated that fae6 was thermolabile with a half life ($T_{1/2}$) < 30 min at $50^{\circ}C$. High affinity for Fae6 against methyl sinapate, methyl ferulate and ethyl ferulate suggest that the enzyme can be useful in hydrolyzing ferulated polysaccharides in a biorefinery process.

• Journal of Microbiology and Biotechnology
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• v.19 no.2
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• pp.187-193
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• 2009

A metagenomic library of surface-water microbes from the Yangtze River in China was constructed, and a novel esterase, designated as EstY, was isolated and characterized. EstY had 423 amino acids with an estimated molecular mass of 44 kDa and pI of 7.28. It hydrolyzed various p-nitrophenyl esters(acetate, butyrate, caprate, caprylate, laurate, myristate, and palmitate) and its best substrate was p-nitrophenyl caprate(C8). The optimum pH for EstY activity was 9.0 and the optimum temperature was $50^{\circ}C$. Metal ions, such as $Mn^{2+},\;Co^{2+},\;Hg^{2+},\;Zn^{2+},\;and\;Fe^{3+}$, strongly inhibited the activity of EstY, whereas $Mg^{2+}$ was required for maximal activity. Activity remained in the presence of 10% alcohol, acetone, isopropanol, and dimethyl sulfoxide, respectively. An analysis of the amino acid sequence deduced from estY revealed that it had 7 closely related lipolytic enzymes. Moreover, a sequence analysis showed that EstY, like its 7 relatives, did not belong to any known lipolytic enzyme family.