Cloning of an epoxide hydrolase‐encoding gene from <i>Rhodotorula mucilaginosa</i> and functional expression in <i>Yarrowia lipolytica</i>

Labuschagne, M. T.; Albertyn, Jacobus

doi:10.1002/yea.1437

Cited by 25 publications

(19 citation statements)

References 36 publications

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“…21 All isolates were strictly aerobic, Gram-negative rods with yellow flexirubin or carotenoid pigments. All isolates were positive for the production of catalase, oxidase, phosphatase, DNase; growth at 25 °C and growth in 0 and 1% NaCl.…”

Section: Identification Of Isolatesmentioning

confidence: 97%

“…21 Polymerase chain reaction (PCR) amplification of the 16S rRNA gene was performed using an Eppendorf Mastercycler Temperature Gradient Personal Thermal cycler (Applied Biosystems, Johannesburg, South Africa) and the forward, 27F (5'-GAGTTTGATCCTGGCTCAG-3') and reverse, 1492R (5'-GGTTACCTTGTTACGACTT-3') primers. 22 The PCR product was visualised on a 1% (w/v) agarose gel.…”

Section: S Rrna Sequencingmentioning

confidence: 99%

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Spoilage potential of a novel group of bacteria isolated from dairy products

Tsôeu¹,

Jooste²,

Charimba³

et al. 2016

S. Afr. J. Sci.

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Cold-tolerant bacteria, also known as psychrotrophic bacteria, are notorious contaminants of milk in the refrigerated dairy food chain. These organisms, especially the pseudomonads, may produce heat-resistant enzymes that are responsible for the breakdown of proteins and lipids in milk and dairy products. Such reactions result in a variety of defects in the raw or unprocessed milk that may affect the suitability of such milk for further processing. The enzymes produced may cause defects in long-life dairy products such as cheese, butter and long-life milk. In the present study, a range of 18 yellow pigmented psychrotrophic bacteria, collectively known as flavobacteria, were isolated from local dairy products. One aim of this study was to identify these bacteria to species level using molecular techniques. A second aim was to determine the spoilage potential of these organisms based on profiles generated by the BIOLOG system (that may relate to hydrolytic enzymes produced). Of the 18 isolates, 14 belonged to the genus Chryseobacterium while 4 were identified as Empedobacter isolates. The most active spoilage organisms in this group were shown to be C. bovis, C. shigense and E. brevis. These findings illustrate that enzymatically catalysed defects in dairy products should not be attributed solely to acknowledged psychrotrophic bacteria such as the pseudomonads, but that flavobacterial species may also be actively involved.

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Section: Identification Of Isolatesmentioning

confidence: 97%

Section: S Rrna Sequencingmentioning

confidence: 99%

Spoilage potential of a novel group of bacteria isolated from dairy products

Tsôeu¹,

Jooste²,

Charimba³

et al. 2016

S. Afr. J. Sci.

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“…In cells, EHs can play important roles such as detoxification agents for epoxide derivatives, for the biosynthesis of signal molecules and the utilization of epoxides as carbon source [2,37]. EHs are ubiquitous in nature and have been found in bacteria [15,16], yeasts [17,40], filamentous fungi [21,27], plants [1,22], insects [33,41], mammals [3] and human tissues [11,18].…”

Section: Introductionmentioning

confidence: 99%

Production of epoxide hydrolases in batch fermentations of Botryosphaeria rhodina

Melzer

Junne

Wohlgemuth

et al. 2008

J Ind Microbiol Biotechnol

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The filamentous fungus Botryosphaeria rhodina (ATCC 9055) was investigated related to its ability for epoxide hydrolase (EH) production. Epoxide hydrolase activity is located at two different sites of the cells. The larger part is present in the cytosol (70%), while the smaller part is associated to membranes (30%). In media optimization experiments, an activity of 3.5 U/gDW for aromatic epoxide hydrolysis of para-nitro-styrene oxide (pNSO) could be obtained. Activity increased by 30% when pNSO was added to the culture during exponential growth. An increase of enzyme activity up to 6 U/gDW was achieved during batch-fermentations in a bioreactor with 2.7 l working volume. Evaluation of fermentations with 30 l working volume revealed a relation of oxygen uptake rate to EH expression. Oxygen limitation resulted in a decreased EH activity. Parameter estimation by the linearization method of Hanes yielded Km values of 2.54 and 1.00 mM for the substrates S-pNSO and R-pNSO, respectively. vmax was 3.4 times higher when using R-pNSO. A protein purification strategy leading to a 47-fold increase in specific activity (940 U/mgProtein) was developed as a first step to investigate molecular and structural characteristics of the EH.

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“…Epoxide hydrolases have been found in a variety of organisms and are thought to be ubiquitous in nature (Weijers and de Bont 1999; Visser et al. 2002; Labuschagne and Albertyn 2007). This coupled with the commercial importance of EH has led to its expression in various recombinant hosts, including Yarrowia lipolytica (Visser et al.…”

Section: Introductionmentioning

confidence: 99%

“…This coupled with the commercial importance of EH has led to its expression in various recombinant hosts, including Yarrowia lipolytica (Visser et al. 2002; Labuschagne and Albertyn 2007; Maharajh et al. 2008).…”

Section: Introductionmentioning

confidence: 99%

Effect of an exponential feeding regime on the production ofRhodotorula araucariaeepoxide hydrolase inYarrowia lipolytica

Maharajh¹,

Lalloo²,

Görgens

2008

Letters in Applied Microbiology

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Aims: To evaluate the effect of and exponential feeding regime on the production of epoxide hydrolase (EH) enzyme in recombinant Yarrowia lipolytica in comparison to a constant feed strategy. Methods and Results: An exponential feed model was developed and fermentations were fed at six different exponential rates. A twofold increase in EH productivity and a 15% increase in volumetric EH activity was obtained by applying exponential glucose feed rates in fed‐batch cultivation. These responses were modelled to obtain a theoretical optimum feed rate that was validated in duplicate fermentations. The model optimum of 0·06 h−1 resulted in a volumetric EH activity of c. 5500 U l−1 h−1 and a maximum activity of 206 000 U l−1. This correlated well with model predictions, with a variance of <10%. Conclusions: The use of an exponential feed strategy at a rate of 0·06 h−1 yielded best results for all key responses which show a clear improvement over a constant feed strategy. Significance and Impact of the Study: The study was the first evaluation of an exponential feed strategy on recombinant Y. lipolytica for the production of EH enzyme. The results suggest a strategy for the commercial production of a valuable pharmaceutical enzyme.

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Cloning of an epoxide hydrolase‐encoding gene from Rhodotorula mucilaginosa and functional expression in Yarrowia lipolytica

Cited by 25 publications

References 36 publications

Spoilage potential of a novel group of bacteria isolated from dairy products

Spoilage potential of a novel group of bacteria isolated from dairy products

Production of epoxide hydrolases in batch fermentations of Botryosphaeria rhodina

Effect of an exponential feeding regime on the production ofRhodotorula araucariaeepoxide hydrolase inYarrowia lipolytica

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