Development of an asparagine-reducing yeast by adaptive evolution and uses thereof to reduce acrylamide formation
Inventors
Turgeon, Zachari J. • Swanson, Jessica Marie • Dahabieh, Matthew S. • Husnik, John I.
Assignees
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Publication Number
US-11051535-B2
Publication Date
2021-07-06
Expiration Date
Abstract
The present disclosure relates to a method of isolating a yeast strain that is able to degrade L-asparagine under non-inducing conditions comprising repeated cycles of adaptive evolution and mutagenesis followed by strain selection. Also included are yeast strains obtained by the method, and methods and uses thereof for reducing asparagine, and thus acrylamide, during food preparation and processing.
Core Innovation
The invention relates to an isolated industrial yeast that degrades L-asparagine under non-inducing conditions. The approach aims to achieve constitutive L-asparagine degradation without genetically modifying the yeast, and links L-asparagine degradation to cell-wall associated Asparaginase II (ASP3/encoded ASP3).
The description addresses the problem that L-asparagine can lead to acrylamide formation in foods and seeks yeast that degrades L-asparagine under non-inducing conditions, including conditions that do not induce nitrogen catabolite repression (NCR), while maintaining relevant Asparaginase activity. It describes selection and enrichment for yeast with sustained asparagine degradation behavior in rich media and non-inducing/non-NCR conditions.
The description presents iterative adaptive evolution combined with mutagenesis and selection under specific selective pressure, resulting in multiple random mutations relative to cells prior to subculturing. The resulting yeast shows constitutive expression of ASP3 while keeping ASP1 expression unchanged, alongside global gene-expression shifts involving NCR genes and amino-acid transporters.
Claims Coverage
The patent includes four independent claims covering a method for producing an isolated industrial yeast strain and three isolated industrial yeast strains deposited with IDAC under accession numbers 140515-01 (RBAR-01), 140515-02 (RBAR-02), and 140515-03 (RBAR-03), each characterized by degrading L-asparagine under non-inducing conditions. The independent claims collectively cover both the strain-production method and three deposited yeast isolates with the same functional activity.
Isolated industrial yeast with non-inducing L-asparagine degradation via iterative mutagenesis and methylamine selection
An isolated industrial yeast that degrades L-asparagine under non-inducing conditions produced by iterative subculturing, mutagenesis, growth-rate tracking, selection in methylamine-containing selective media, colony isolation, assaying for L-asparagine degradation under non-inducing conditions, and repeated selection until activity reaches a plateau.
IDAC-deposited RBAR-01 isolate degrading L-asparagine under non-inducing conditions
An isolated industrial yeast strain deposited with the International Depositary Authority of Canada (IDAC) under accession number 140515-01 (RBAR-01) wherein the yeast degrades L-asparagine under non-inducing conditions.
IDAC-deposited RBAR-02 isolate degrading L-asparagine under non-inducing conditions
An isolated industrial yeast strain deposited with the International Depositary Authority of Canada (IDAC) under accession number 140515-02 (RBAR-02) wherein the yeast degrades L-asparagine under non-inducing conditions.
IDAC-deposited RBAR-03 isolate degrading L-asparagine under non-inducing conditions
An isolated industrial yeast strain deposited with the International Depositary Authority of Canada (IDAC) under accession number 140515-03 (RBAR-03) wherein the yeast degrades L-asparagine under non-inducing conditions.
The central inventive concept is an isolated industrial yeast characterized by degrading L-asparagine under non-inducing conditions, including a method for producing such yeast via iterative adaptive evolution and mutagenesis with methylamine selection, and three specific IDAC-deposited RBAR isolates defined by the same functional activity.
Stated Advantages
Reduces asparagine and/or acrylamide concentrations in food products by including the claimed yeast strain.
Documented Applications
Food products, including bread, toast, French fries/snacks, sweet biscuits, pretzels, and coffee/green coffee beans/green coffee extract, treated using the yeast to show reduced acrylamide formation and associated L-asparagine depletion.
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