Double mutation of Saccharomyces cerevisiae for enhanced β-D-fructofuranosidase fructohydrolase productivity and application of growth kinetics for parametric significance analysis

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Abstract

The kinetics of an extracellular β-d-fructofuranosidase fructohydrolase production by Saccharomyces cerevisiae in a chemically defined medium, i.e., sucrose peptone agar yeast extract at pH 6, was investigated. The wild-type was treated with a chemical mutagen, methyl methane sulfonate. Among the six mutants isolated, methyl methane sulfonate-V was found to be a better enzyme producing strain (52 ± 2.4aU/mL). The maximum production (74 ± 3.1aU/mL) was accomplished after at 48 h (68 ± 2.7amg/mL protein). The mutants were stabilized at low levels of 5-fluoro-cytocine and the viable ones were further processed for optimization of cultural conditions and nutritional requirements. The sucrose concentration, incubation period and pH were optimized to be 30 g/L, 28°C, and 6.5, respectively. The methyl methane sulfonate-V exhibited an improvement of over 10 folds in enzyme production when 5 g/L ammonium sulfate was used as a nitrogen source. Thin layer chromatography and high-performance liquid chromatography analysis illustrated the optimalenzyme activity supported by the higher rate of hydrolysis of sucrose into monosaccharides, particularly α-d-glucose and β-d-fructose. The values for Qp(2 ± 0.12cU/mL/h) and Yp/s(4 ± 1.24bU/g) of the mutant were considerably increased in comparison with other yeast strains (both isolates and viable mutants). The mutant could be exploited for enzyme production over a wider temperature range (26–34°C), with significantly high enzyme activity (LSD 0.048, HS) at the optimal temperature.

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Ali, S., Aslam, A., & Hayyat, M. U. (2016). Double mutation of Saccharomyces cerevisiae for enhanced β-D-fructofuranosidase fructohydrolase productivity and application of growth kinetics for parametric significance analysis. Brazilian Journal of Microbiology, 47(1), 136–142. https://doi.org/10.1016/j.bjm.2015.11.024

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