Methods and strains for producing bioproducts in Aureobasidium pullulans
Inventors
Skory, Christopher D. • Leathers, Timothy D. • Price, Neil P
Assignees
US Department of Agriculture USDA
Publication Number
US-10351889-B2
Publication Date
2019-07-16
Expiration Date
2036-07-18
Interested in licensing this patent?
MTEC can help explore whether this patent might be available for licensing for your application.
Abstract
The present disclosure provides methods for producing bioproducts from novel genetically altered strains of Aureobasidium pullulans. Methods and materials for the construction of these strains, examination of the bioproducts and analysis and isolation of the bioproducts from genetically altered strains is provided. Genetically altered A. pullulans strains in which one or more genes encoding biosynthetic enzymes are knocked out is detailed and the benefits of using such strains described.
Core Innovation
The invention provides methods and novel genetically altered strains of Aureobasidium pullulans for producing bioproducts. These genetically altered strains have one or more genes encoding biosynthetic enzymes knocked out, such as mannitol-1-phosphate-dehydrogenase (MPD1), mannitol-dehydrogenase (MDH2), and polyketide synthase (PKS) genes. By doing so, the strains preferentially produce target bioproducts like liamocins, pullulan, and massoia lactone, among others. The disclosure also provides methods for constructing, examining, analyzing, and isolating bioproducts from these genetically modified strains.
The problem addressed involves the production of undesirable bio-contaminants such as melanin and related pigments by wild-type A. pullulans strains. These pigments contaminate bioproducts, requiring costly purification steps including activated charcoal or hydrogen peroxide treatment, ultrafiltration, and ion exchange resins. Such purification results in losses of the desired bioproduct and increased manufacturing costs. Additionally, non-specific production of other undesirable bioproducts lowers overall yield and economic feasibility.
The invention solves these problems by genetically altering A. pullulans strains to lack functional biosynthetic enzymes responsible for melanin and undesired products, enabling direct production of substantially melanin-free bioproducts. For example, strains lacking a functional PKS gene do not produce melanin, resulting in melanin-free bioproducts like pullulan, liamocins, massoia lactone, exophilins, poly(β-malic acid), β-glucan, aureobasidin, intracellular fatty acids, and triacylglycerols. Strains lacking MPD1 or MDH2 genes can be used to produce arabitol-liamocins on inexpensive carbon sources such as glucose or fructose, avoiding the prior requirement for costly substrates.
Claims Coverage
The patent contains one independent claim defining a broad method for producing a melanin-free bioproduct utilizing genetically altered Aureobasidium pullulans strains.
Method of producing melanin-free bioproducts with polyketide synthase gene disruption
A method of producing a melanin-free bioproduct selected from pullulan, liamocins, lactones, exophilins, poly(β-malic acid), β-glucan, aureobasidin, intracellular fatty acids, and triacylglycerols by growing an A. pullulans strain lacking a functional polyketide synthase (PKS4) gene under conditions supporting bioproduct production, followed by collection of the bioproduct.
Use of a specific genetically altered A. pullulans strain for melanin-free bioproduct production
Utilization of a genetically pure A. pullulans strain deposited as NRRL 67080 characterized by lacking a functional PKS4 gene to produce melanin-free bioproducts as disclosed in the method above.
The claims focus on methods of producing melanin-free bioproducts by using A. pullulans strains genetically modified to lack a functional PKS4 gene, including the use of particular deposited strains, enabling production of various bioproducts without melanin contamination.
Stated Advantages
Reduction or elimination of melanin contamination in bioproducts produced by A. pullulans.
Lower purification costs due to melanin-free bioproducts requiring fewer or no downstream purification steps to remove pigments.
Increased economic feasibility and value of bioproduct production from genetically modified strains on cheaper substrates such as glucose instead of more costly carbon sources.
Ability to preferentially produce desired bioproducts while reducing or eliminating production of undesired bioproducts.
Provision of more environmentally responsible and cost-effective production methods for food, pharmaceutical, industrial, and biofuel applications.
Documented Applications
Production of pullulan for food, pharmaceutical, and industrial applications such as emulsifiers, thickeners, edible films, coatings, and packaging.
Production of liamocins exhibiting antibacterial and biosurfactant properties for pharmaceutical, veterinary, and industrial uses.
Production of massoia lactone as a natural coconut flavoring ingredient and potential applications in artificial sweeteners and fuel additives.
Production of other bioproducts such as exophilins, poly(β-malic acid), β-glucan, aureobasidin, intracellular fatty acids, and triacylglycerols.
Interested in licensing this patent?