Summary: The methylotrophic yeast was grown in a chemostat with a medium containing a mixture of glucose (C) and methanol (C) (87·8% C: 12·2% C, w/w) as sole carbon source and NH as sole nitrogen source. At a constant growth rate ( 0·10 h) the influence of the carbon: nitrogen ratio (C:N) of the inflowing medium on the cellular and enzymic composition of the cells was studied. Three distinct growth regimes were recognized. A medium with a C:N ratio < 12 resulted in carbon-limited growth (high cellular protein content, low carbohydrate content) and under these conditions glucose and methanol were utilized simultaneously. A medium with a C:N ratio>31 resulted in nitrogen-limited growth (low protein but high carbohydrate content of the cells) and the cells metabolized only glucose. A transition growth regime was observed during growth on media with intermediate C:N ratios (12 < C:N> 31). When assessed from both substrate consumption and cellular composition, growth was double-substrate (carbon and nitrogen)-limited. In this transition growth regime, changes in carbon metabolism and the cellular and enzymic composition of the cells were found. With increasing C:N ratios in the growth medium a gradual repression of the synthesis of methanol-assimilating and dissimilating enzymes was found. This effect was most pronounced for alcohol oxidase, and as a consequence the cells switched from the utilization of the carbon substrate mixture to growth on glucose alone. The data presented suggest that the range within which double-substrate-limited growth can be expected is predictable from the composition of cells grown under single substrate limitation.


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