Activities of Regulatory Enzymes in Alkane-utilizing and Lipid-accumulating Yeasts and Moulds

Whitworth, D. A.; Ratledge, Colin

doi:10.1099/00221287-90-1-183

Cited by 4 publications

(1 citation statement)

References 8 publications

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“…Under these circumstances, utilization of carbon and nitrogen was less efficient and both biomass and lipid yields were poor. The factors causing lipid to accumulate may therefore be different with different limiting nutrients, but to substantiate this a firmer understanding of the biochemical basis of lipid accumulation in oleaginous organisms is needed (25,26).…”

Section: Consistencymentioning

confidence: 99%

Lipid accumulation in an oleaginous yeast (Candida 107) growing on glucose in single-stage continuous culture

Gill¹,

Hall²,

Ratledge³

1977

Appl Environ Microbiol

153

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Lipid accumulation in Candida 107, grown at dilution rates from 0.03 to the maximum of 0.21/h, with carbon, nitrogen, phosphate, and magnesium limitations in a chemostat, was maximal at about 40% (wt/wt) with nitrogen-limited medium at a dilution rate of 0.06/h, giving an efficiency of substrate conversion of 22 g of lipid per 100 g of glucose consumed. At higher dilution rates the lipid content decreased. With carbon-limited growth, the highest lipid content (14%, wt/wt) was at the maximum dilution rate. High lipid contents also occurred with phosphate + nitrogen as double limitations of growth, with the lipid content of the yeast (about 35%, wt/wt) continuing to be near maximum at dilution rates also near maximum (0.17/h), thus giving the highest specific rate of lipid formation of any growth conditions (0.059 g of lipid/g of yeast per h). However, the efficiency of substrate utilization was only 5.2 g of lipid formed per 100 g of glucose consumed. The composition of the fatty acyl residues within the lipid remained constant over many weeks if the steady-state conditions remained unchanged. With carbon-limited growth, the degree of unsaturation of the fatty acids markedly decreased as the dilution rate was increased, but with nitrogen limitation the reverse trend was seen. In all cases, linoleic and oleic acids were the principal fatty acyl residues affected, and their relative proportions always varied in opposite directions. When magnesium was a limiting nutrient, there was a considerable increase in the proportion of myristic acid produced within the lipid. Neutral lipids (predominantly triglycerides) varied from 66 to 92% of the total lipid from carbonand nitrogen-limited growth; phospholipids (varying from 2 to 25%) were highest in nitrogen-limited growth. The fatty acyl residues within each lipid fraction showed the same variations with changing growth rates.

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Section: Consistencymentioning

confidence: 99%

Lipid accumulation in an oleaginous yeast (Candida 107) growing on glucose in single-stage continuous culture

Gill¹,

Hall²,

Ratledge³

1977

Appl Environ Microbiol

153

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Lipid biosynthesis in oleaginous yeasts

Turcotte

Kosaric

Advances in Biochemical Engineering/Biotechnology

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Pathways of Glucose Metabolism in Candida 107, a Lipid-accumulating Yeast

Ratledge

Botham

1977

Journal of General Microbiology

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Phosphofructokinase was not detected in extracts of Candida 107 prepared in a variety of ways but was highly active in cells treated with toluene. Disruption of these cells destroyed activity of phosphofructokinase indicating that the enzyme is extremely labile. As patterns of labelling from [~-~~C]glucose and [6-14C]glucose showed that 60 % of glucose was metabolized via the pentose cycle, augmentation of this cycle is necessary to account for the high molar growth yields of this yeast. Phosphoketolases, reacting with xylulose 5-phosphate and fructose 6-phosphate, were found but the extent to which they contribute to glucose metabolism was not assessed. I N T R O D U C T I O NWe have previously examined the intermediary metabolism of an oleaginous (fatproducing) yeast, Candida 107, and, because of the apparent absence of phosphofructokinase, concluded that glucose may b: metabolized via the pentose cycle (Whitworth & Ratledge, 1975a). Metabolism of glucose by this cycle, although providing NADPH for fatty acid biosynthesis, would be inefficient with only I mol acetyl-CoA arising from I mol glucose. An alternative must exist to explain the high molar growth and lipid yields of this organism (Gill, Hall & Ratledge, 1977). The same conclusion was reached by Hofer and co-workers (Hofer, 1968;Hofer et a/., 1971) and by Brady & Chambliss ( I 967) working with another fat-accumulating yeast, Rhodotorula gracilis (now Rhodosporidium toruloides), in which phosphofructokinase also seemed to be absent. Hofer et a/. (1969) suggested that phosphoketolase may be present to split xylulose 5-phosphate (formed by the transaldolase and transketolase reactions) into glyceraldehyde 3-phosphate and acetyl-CoA but did not look for this enzyme. However, Mazon, Gancedo & Gancedo (1974) found a phosphofructokinase in Rhodotorula glutinis which was active only under carefully controlled conditions, indicating that caution was needed in interpreting the previous results. When we repeated the latter workers' assay with extracts of Candida 107 we still failed to detect this enzyme and therefore began an examination of glucose metabolism in this yeast. Our results are presented here. M E T H O D SOrganisms and growth. Candida 107 Ruinen 8c Deinema 1964 and, as a control organism, Saccharomyces cerevisiae were grown at 30 "C in a medium consisting of (g 1-9: glucose, 12; KH2P04, 7; Na,HPO,, 2 ; NH,CI, 1-7; MgS04.7Ho0, 1.5; yeast extract, 1.5; CaC1,.6H20, 0 . 1 ; FeCI3.6H,O, 0.05; ZnS04.7H,0, 0-0001; with a final pH of 5-5. The yeasts were usually grown in 100 ml medium in 250 ml conical flasks but occasionally in a chemostat as previously described (Gill et al., 1977). Yeasts were harvested at the end of active growth by centrifuging at 10000 g for 5 to 10 min, washed once with 50 mwphosphate buffer, pH 7-5, containing 2 mM-MgC1, and then resuspended in the same buffer solution to about 300 mg dry wt ml-l.Preparation of cell-free extracts. Conventional procedures were followed for disrupting cells (Hughes,

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Activities of Regulatory Enzymes in Alkane-utilizing and Lipid-accumulating Yeasts and Moulds

Cited by 4 publications

References 8 publications

Lipid accumulation in an oleaginous yeast (Candida 107) growing on glucose in single-stage continuous culture

Lipid accumulation in an oleaginous yeast (Candida 107) growing on glucose in single-stage continuous culture

Lipid biosynthesis in oleaginous yeasts

Pathways of Glucose Metabolism in Candida 107, a Lipid-accumulating Yeast

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