Small Differences in Gene Sequences Impact Invertase Activity and Specificity toward Fructans with Different Chain Lengths.

invertase is encoded by a family of closely related genes. To identify and understand the molecular basis for differences in substrate specificity, we examined 29 alleles from industrialstrains and cloned alleles with small sequence differences into an invertase-negative strain. Our study showed that an F102Y substitution in Suc-enzymes lowers yeast invertase activity toward fructo-oligosaccharides (FOS) by 36% and the specificity factor by 43%. By contrast, an A409P substitution in Suc-enzymes resulted in an increased capacity of the yeast to hydrolyze FOS and Fibruline by 17 and 41%, respectively, likely because of a change in the loop conformation resulting in a wider active site. Bread dough fermentation experiments revealed that sucrose and fructan hydrolysis during fermentation is influenced by this natural variation in sequences. Our research thus opens the door for the selection or engineering of yeasts and Suc-enzymes with specific activities that may ultimately allow controlling fructan hydrolysis.