• Microbiology and Biotechnology Letters
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• v.24 no.3
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• pp.316-320
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• 1996

An alkaline proteinase produced by Yarrowia lipolytica TH65 was purified by 40-65% ammonium sulfate fractionation, DEAE-cellulose chromatography, and gel filtration with Sephadex G-100 and Sephadex G-75. The purified enzyme was shown as a single band on SDS-PAGE, and its molecular weight 31,500. Optimum temperature and pH were 40$\circ$C and 8.5-9.0, respectively, and the enzyme was stable below 40$\circ$C and in the pH range of 6-8. The enzyme was strongly inhibited by divalent ions, completely by PMSF, and partially by EDTA, EGTA, and phenanthroline. But the inhibitory effect in the presence of EDTA, EGTA and phenanthroline could be reversed by addition of Ca$^{2+}$. Thus, these results indicated that the purified enzyme was an alkaline serine proteinase (E.C. 3.4.21.14).

• Journal of Applied Biological Chemistry
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• v.50 no.2
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• pp.46-51
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• 2007

Lipases are industrially useful versatile enzymes that catalyze numerous different reactions. Among lipases functioning under extreme conditions, alkaline lipase is useful in detergent industry. Lipase from yeast strain Yarrowia lipolytica NRRL Y-2178 was most active under alkaline condition, and initial medium pH for most lipase production was also alkaline [Lee et al., 2007, J Microbiol Biotechnol, 17(6)]. High lipase production was achieved using Y. lipolytica NRRL Y-2178. Optimal incubation time for lipase production at $25^{\circ}C$ was 72 h. Optimal temperature, when incubated for 72 h, was $27.5^{\circ}C$. Lipase production but not cell growth was very sensitive to concentrations of glucose and glycerol as efficient carbon sources, showing optimal concentrations of 1.0 and 1.5% (w/v), respectively. Lipase production was highly stimulated by $Ca^{2+},\;K^+,\;and\;Na^+$, but was inhibited by $Co^{2+},\;Cu^{2+},\;Mn^{2+},\;Na^+,\;and\;Fe^{2+}$. Maximum lipase production at 0.1 mM $Ca^{2+}$ for 72 h incubation at $27.5^{\circ}C$ was 649 units/mL.

• Journal of Microbiology and Biotechnology
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• v.28 no.4
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• pp.597-605
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• 2018

Acyl-CoA oxidases (ACOXs) play important roles in lipid metabolism, including peroxisomal fatty acid ${\beta}$-oxidation by the conversion of acyl-CoAs to 2-trans-enoyl-CoAs. The yeast Yarrowia lipolytica can utilize fatty acids as a carbon source and thus has extensive biotechnological applications. The crystal structure of ACOX3 from Y. lipolytica (YlACOX3) was determined at a resolution of $2.5{\AA}$. It contained two molecules per asymmetric unit, and the monomeric structure was folded into four domains; $N{\alpha}$, $N{\beta}$, $C{\alpha}1$, and $C{\alpha}2$ domains. The cofactor flavin adenine dinucleotide was bound in the dimer interface. The substrate-binding pocket was located near the cofactor, and formed at the interface between the $N{\alpha}$, $N{\beta}$, and $C{\alpha}1$ domains. Comparisons with other ACOX structures provided structural insights into how YlACOX has a substrate preference for short-chain acyl-CoA. In addition, the structure of YlACOX3 was compared with those of medium- and long-chain ACOXs, and the structural basis for their differences in substrate specificity was discussed.

• Journal of Microbiology
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• v.37 no.3
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• pp.128-135
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• 1999

Among oil-degrading microorganisms isolated from oil-polluted industrial areas, one yeast strain showed high degradation activity of aliphatic hydrocarbons. From the analyses of 18S rRNA sequences, fatty acid, coenzyme Q system, G+C content of DNA, and biochemical characteristics, the strain was identified as Yarrowia lipolytica 180. Y. lipolytica 180 degraded 94% of aliphatic hydrocarbons in minimal salts medium containing 0.2% (v/v) of Arabian light crude oil within 3 days at 25$^{\circ}C$. Optimal growth conditions for temperature, pH, NaCl concentration, and crude oil concentration were 30$^{\circ}C$, pH 5-7, 1%, and 2% (v/v), respectively. Y. lipolytica 180 reduced surface tension when cultured on hydrocarbon substrates (1%, v/v), and the measured values of the surface tension were in the range of 51 to 57 dynes/cm. Both the cell free culture broth and cell debris of Y. lipolytica 180 were capable of emulsifying 2% (v/v) crude oil by itself. They were also capable of degrading crude oil (2%). The strain showed a cell surface hydrophobicity higher than 90%, which did not require hydrocarbon substrates for its induction. These results suggest that Y. lipolytica has high oil-degrading activity through its high emulsifying activity and cell hydrophobicity, and further indicate that the cell surface is responsible for the metabolism of aliphatic hydrocarbons.

• Journal of Microbiology and Biotechnology
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• v.17 no.6
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• pp.1054-1057
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• 2007

Lipases are industrially useful versatile enzymes that catalyze numerous different reactions including hydrolysis of triglycerides, transesterification, and chiral synthesis of esters under natural conditions. Although lipases from various sources have been widely used in industrial applications, such as in food, chemical, pharmaceutical, and detergent industries, there are still substantial current interests in developing new microbial lipases, specifically those functioning in abnormal conditions. We screened 17 lipase-producing yeast strains, which were prescreened for substrate specificity of lipase from more than 500 yeast strains from the Agricultural Research Service Culture Collection (Peoria, IL, U.S.A.), and selected Yarrowia lipolytica NRRL Y-2178 as a best lipase producer. This report presents new finding and optimal production of a novel extracellular alkaline lipase from Y. lipolytica NRRL Y-2178. Optimal culture conditions for lipase production by Y. lipolytica NRRL Y-2178 were 72 h incubation time, $27.5^{\circ}C$, pH 9.0. Glycerol and glucose were efficiently used as the most efficient carbon sources, and a combination of yeast extract and peptone was a good nitrogen source for lipase production by Y. lipolytica NRRL Y-2178. These results suggested that Y. lipolytica NRRL Y-2178 shows good industrial potential as a new alkaline lipase producer.

• Biotechnology and Bioprocess Engineering:BBE
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• v.5 no.5
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• pp.320-326
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• 2000

Promoters of the genes G3P, ICL1, POT1, POX1, POX2 and POX5 of the yeast Y. lipolytica were studied in respect to their regulations and activities during growth on different carbon sources. The aim of this study was to select suitable promoters for high expression of heterologous genes in this yeast. For this purpose the promoters were fused with the reporter gene lacZ of E. coli and integrated as single copies into the genome of Y. lipolytica strain PO1d. The measurement of expressed activities of ${\beta}$-galactosidase revealed that pICL1, pPOX2 and pPOT1 are the strongest regulable promoters available for Y. lipolytica, at present. pPOX2 and pPOT1 were highly induced during growth on oleic acid and were completely repressed by glucose and glycerol. pICL1 was strongly inducible by ethanol besides alkanes and fatty acids, however, not completely repressible by glucose or glycerol. Ricinoleic acid methyl ester appeared as a very strong inducer for pPOT1 and pPOX2, in spite of that it inhibited growth of Y. lipolytica transformants.

• Journal of Microbiology and Biotechnology
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• v.25 no.7
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• pp.1056-1069
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• 2015

Osmotic pressure is a critical factor for erythritol production with osmophilic yeast. Protein expression patterns of an erythritol-producing yeast, Yarrowia lipolytica, were analyzed to identify differentially-expressed proteins in response to osmotic pressure. In order to analyze intracellular protein levels quantitatively, two-dimensional gel electrophoresis was performed to separate and visualize the differential expression of the intracellular proteins extracted from Y. lipolytica cultured under low (3.17 osmol/kg) and high (4.21 osmol/kg) osmotic pressures. Proteomic analyses allowed identification of 54 differentially-expressed proteins among the proteins distributed in the range of pI 3-10 and 14.4-97.4 kDa molecular mass between the osmotic stress conditions. Remarkably, the main proteins were involved in the pathway of energy, metabolism, cell rescue, and stress response. The expression of such enzymes related to protein and nucleotide biosynthesis was inhibited drastically, reflecting the growth arrest of Y. lipolytica under hyperosmotic stress. The improvement of erythritol production under high osmotic stress was due to the significant induction of a range of crucial enzymes related to polyols biosynthesis, such as transketolase and triosephosphate isomerase, and the osmotic stress responsive proteins like pyridoxine-4-dehydrogenase and the AKRs family. The polyols biosynthesis was really related to an osmotic response and a protection mechanism against hyperosmotic stress in Y. lipolytica. Additionally, the high osmotic stress could also induce other cell stress responses as with heat shock and oxidation stress responses, and these responsive proteins, such as the HSPs family, catalase T, and superoxide dismutase, also had drastically increased expression levels under hyperosmotic pressure.

• Animal Bioscience
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• v.35 no.4
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• pp.605-613
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• 2022

Objective: The objective was to evaluate the efficacy of increasing supplementation of Yarrowia lipolytica (YL) up to 3.0% replacing 1.6% poultry fat and 0.9% blood plasma for growth performance, intestinal health and nutrient digestibility of diets fed to nursery pigs. Methods: Twenty-four pigs weaned at 24 d of age (initial body weight at 7.2±0.6 kg) were allotted to three dietary treatments (n = 8) based on the randomized complete block. The diets with supplementation of YL (0.0%, 1.5%, and 3.0%, replacing poultry fat and blood plasma up to 1.6% and 0.9%, respectively) were fed for 21 d. Feed intake and body weight were recorded at d 0, 10, and 21. Fecal score was recorded at every odd day from d 3 to 19. Pigs were euthanized on d 21 to collect proximal and distal jejunal mucosa to measure intestinal health markers including tumor necrosis factor-alpha, interleukin-8, immunoglobulin A and immunoglobulin G. Ileal digesta was collected for apparent ileal digestibility (AID) of nutrients in diets. Data were analyzed using Proc Mixed of SAS. Results: Supplementation of YL (1.5% and 3.0%) replacing poultry fat and blood plasma did not affect growth performance, fecal score and intestinal health. Supplementation of YL at 1.5% did not affect nutrient digestibility, whereas supplementation of YL at 3.0% reduced AID of dry matter (40.2% to 55.0%), gross energy (44.0% to 57.5%), crude protein (52.1% to 66.1%), and ether extract (50.8% to 66.9%) compared to diets without supplementation. Conclusion: Yarrowia lipolytica can be supplemented at 1.5% in nursery diets, replacing 0.8% poultry fat and 0.45% blood plasma without affecting growth performance, intestinal health and nutrient digestibility. Supplementation of YL at 3.0% replacing 1.6% poultry fat and 0.9% blood plasma did not affect growth performance and intestinal health, whereas nutrient digestibility was reduced.

• Journal of Microbiology and Biotechnology
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• v.18 no.9
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• pp.1522-1528
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• 2008

The morphogenetic behavior of a tropical marine Yarrowia lipolytica strain on hydrophobic substrates was studied. Media containing coconut oil or palm kernel oil (rich in lauric and myristic acids) prepared in distilled water or seawater at a neutral pH supported 95% of the cells to undergo a transition from the yeast form to the mycelium form. With potassium laurate, 51 % of the cells were in the mycelium form, whereas with myristate, 32% were in the mycelium form. However, combinations of these two fatty acids in proportions that are present in coconut oil or palm kernel oil enhanced the mycelium formation to 65%. The culture also produced extracellular lipases during the morphogenetic change. The yeast cells were found to attach to the large droplets of the hydrophobic substrates during the transition, while the mycelia were associated with the aqueous phase. The alkane-grown yeast partitioned more efficiently in the hydrophobic phases when compared with the coconut oil-grown mycelia. A fatty acid analysis of the mycelial form revealed the presence of lauric acid in addition to the long-chain saturated and unsaturated fatty acids observed in the yeast form. The mycelia underwent a rapid transition to the yeast form with n-dodecane, a medium-chain aliphatic hydrocarbon. Thus, the fungus displayed a differential behavior towards the two types of saturated hydrophobic substrates.

• Journal of Microbiology
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• v.40 no.2
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• pp.173-177
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• 2002

In order to identify Yawowia lipolytica genes induced by serum, cDNA representational difference analysis was performed using a PCR-select CDNA subtraction method. One of the genes cloned from the subtraction was a gene (YIMBFl) homologous to Saccharomyces cerevisiae MBF1 encoding the coactivator multiprotein bridging factor 1. Disruption of YIMBFl revealed that the gene was net essential for viability, and the Ylmbf△ strain did not show any distinct phenotypic change on solid serum medium. In liquid medium, however. a difference was found in the ability to maintain hyphae induced by serum. This result suggests that the YIMbf1 protein may mediate transcriptional activation of certain genes involved in the hypha fonmation of Y. lipolytica.