In hex2 mutants of Saccharomyces cerevisiae, which are defective in glucose repression of several enzymes, growth is inhibited if maltose is present in the medium. After adding [14C]maltose to cultures growing with ethanol, maltose metabolism was followed in both hex2 mutant and wild-type cells. The amount of radioactivity incorporated was much higher in hex2 than in wild-type cells. Most of the radioactivity in hex2 cells was located in the low molecular mass fraction. Pulse-chase experiments showed that 2 h after addition of maltose, hex2 cells hydrolysed maltose to glucose, which was partially excreted into the medium. 31P-NMR studies gave evidence that turnover of sugar phosphates was completely abolished in hex2 cells after 2 h incubation with maltose. 13C-NMR spectra confirmed these results: unlike those for the wild-type, no resonances corresponding to fermentation products (ethanol, glycerol) were found for hex2 cells, whereas there were resonances corresponding to glucose. Although maltose is taken up by proton symport, the internal pH in the hex2 mutant did not change markedly during the 5 h after adding maltose. The intracellular accumulation of glucose seems to explain the inhibition of growth by maltose, probably by means of osmotic damage and/or unspecific O-glycosylation of proteins. Neither maltose permease nor maltase was over-expressed, and so these enzymes were not the cause of glucose accumulation. Hence, the coordination of maltose uptake, hydrolysis to glucose and glycolysis of glucose is not regulated simply by the specific activity of the catabolic enzymes involved.(ABSTRACT TRUNCATED AT 250 WORDS)