Co-expression of the A. tubingensis amyA and glaA genes on episomal plasmids conveyed amylolytic activity to both a laboratory (Y294) and a semi-industrial strain (Mnuα1) of S. cerevisiae. The α-amylase and glucoamylase activities reached 1.51 and 1.16 U ml-1, respectively, in the Mnuα1[AmyA-GlaA] strain, which compare favourably with the 96–190 U ml-1 and 140–340 U ml-1 reported previously for α-amylase and glucoamylase expression in other haploid strains [18].
The recombinant S. cerevisiae Mnuα1[AmyA-GlaA] strain was superior in its ability to convert 83% of the available carbon in 20 g l-1 raw corn starch and produced 80% of the theoretical ethanol yield after 10 days. At a higher substrate loading of 200 g l-1 raw corn starch, 61% and 57% of the theoretical ethanol yield was achieved within 5 and 10 days, respectively. The starch was completely hydrolysed (saccharified) with 74% converted to released sugars plus fermentation products (mainly ethanol, glycerol and CO2) and the remainder presumably to yeast biomass. The lower ethanol and residual glucose levels for the S. cerevisiae Y294[AmyA-GlaA] fermentation suggested weaker saccharification by the recombinant S. cerevisiae Y294 strain, whereas fermentation capacity is the limiting factor for the S. cerevisiae Mnuα1[AmyA-GlaA] strain.