Enhancement of the proline and nitric oxide synthetic pathway improves fermentation ability under multiple baking-associated stress conditions in industrial baker's yeast

Abstract

BackgroundDuring the bread-making process, industrial baker's yeast, mostly Saccharomyces cerevisiae, is exposed to baking-associated stresses, such as air-drying and freeze-thaw stress. These baking-associated stresses exert severe injury to yeast cells, mainly due to the generation of reactive oxygen species (ROS), leading to cell death and reduced fermentation ability. Thus, there is a great need for a baker's yeast strain with higher tolerance to baking-associated stresses. Recently, we revealed a novel antioxidative mechanism in a laboratory yeast strain that is involved in stress-induced nitric oxide (NO) synthesis from proline via proline oxidase Put1 and N-acetyltransferase Mpr1. We also found that expression of the proline-feedback inhibition-less sensitive mutant γ-glutamyl kinase (Pro1-I150T) and the thermostable mutant Mpr1-F65L resulted in an enhanced fermentation ability of baker's yeast in bread dough after freeze-thaw stress and air-drying stress, respectively. However, baker's yeast strains with high fermentation ability under multiple baking-associated stresses have not yet been developed.ResultsWe constructed a self-cloned diploid baker's yeast strain with enhanced proline and NO synthesis by expressing Pro1-I150T and Mpr1-F65L in the presence of functional Put1. The engineered strain increased the intracellular NO level in response to air-drying stress, and the strain was tolerant not only to oxidative stress but also to both air-drying and freeze-thaw stresses probably due to the reduced intracellular ROS level. We also showed that the resultant strain retained higher leavening activity in bread dough after air-drying and freeze-thaw stress than that of the wild-type strain. On the other hand, enhanced stress tolerance and fermentation ability did not occur in the put1-deficient strain. This result suggests that NO is synthesized in baker's yeast from proline in response to oxidative stresses that induce ROS generation and that increased NO plays an important role in baking-associated stress tolerance.ConclusionsIn this work, we clarified the importance of Put1- and Mpr1-mediated NO generation from proline to the baking-associated stress tolerance in industrial baker's yeast. We also demonstrated that baker's yeast that enhances the proline and NO synthetic pathway by expressing the Pro1-I150T and Mpr1-F65L variants showed improved fermentation ability under multiple baking-associated stress conditions. From a biotechnological perspective, the enhancement of proline and NO synthesis could be promising for breeding novel baker's yeast strains.

Effects of SNF1 on Maltose Metabolism and Leavening Ability of Baker's Yeast in Lean Dough

“…In the baking industry, further advances such as high fermentation ability, frozen tolerance and osmototolerance of baker's yeasts have been demanded in recent decades to adapt the changed conditions during the baking process (Sasano and others , 2013; Sun and others ; Lin and others ). One of the important technological goals is to develop baker's yeast strains (mostly Saccharomyces cerevisiae ) with high fermentation ability (Alves–Araújo and others ; Sasano and others ).…”

Structural and functional analysis of the yeast N -acetyltransferase Mpr1 involved in oxidative stress tolerance via proline metabolism

“…Because Mpr1 confers AZC resistance on various organisms, selectable marker systems using the MPR1 gene and AZC have been constructed for yeast and plant transformation (33)(34)(35). We recently showed that industrial yeast strains expressing the Mpr1 variant are more tolerant to oxidative stresses, such as ethanol, freezing, and desiccation, than those expressing WT-Mpr1 (29,36,37). Engineered Mpr1 might be useful in breeding oxidative stress-tolerant yeast strains with improved fermentation ability.…”

Natural Flavours Obtained by Microbiological Pathway

“…The proline catabolism by Saccharomyces cerevisiae also leads to flavour biosynthesis, the intermediary compound being glutamate, which is further degraded to aroma compounds [15].…”