Methods and materials for deconstruction of biomass for biofuels production
Inventors
Schoeniger, Joseph S. • Hadi, Masood Zia
Assignees
National Technology and Engineering Solutions of Sandia LLC • Sandia National Laboratories
Publication Number
US-9024111-B1
Publication Date
2015-05-05
Expiration Date
2032-03-30
Interested in licensing this patent?
MTEC can help explore whether this patent might be available for licensing for your application.
Abstract
The present invention relates to nucleic acids, peptides, vectors, cells, and plants useful in the production of biofuels. In certain embodiments, the invention relates to nucleic acid sequences and peptides from extremophile organisms, such as SSO1949 and Ce1A, that are useful for hydrolyzing plant cell wall materials. In further embodiments, the invention relates to modified versions of such sequences that have been optimized for production in one or both of monocot and dicot plants. In other embodiments, the invention provides for targeting peptide production or activity to a certain location within the cell or organism, such as the apoplast. In further embodiments, the invention relates to transformed cells or plants. In additional embodiments, the invention relates to methods of producing biofuel utilizing such nucleic acids, peptides, targeting sequences, vectors, cells, and/or plants.
Core Innovation
The invention relates to nucleic acids, peptides, vectors, cells, and plants useful in the production of biofuels. Specifically, it involves nucleic acid sequences and peptides derived from extremophile organisms, such as SSO1949 and Ce1A, which are effective in hydrolyzing plant cell wall materials. The invention also includes modified versions of these sequences optimized for production in monocot and dicot plants, and methods to target peptide production or activity to specific cellular locations such as the apoplast. Additionally, transformed cells and plants expressing these sequences and methods of producing biofuel using these components are disclosed.
The problem addressed is the inefficiency of existing biofuel production methods from lignocellulosic biomass, due to the recalcitrance of the biomass to chemical pretreatment and enzymatic hydrolysis steps. Current methods suffer from low activity of recombinant deconstruction enzymes, mass transfer issues, high costs, and energy-intensive pretreatment. There is a need for improved methods for biofuel production, particularly methods that improve the breakdown of plant cell wall polysaccharides and optimize enzyme activity.
The invention proposes the introduction of exogenous nucleic acids encoding cellulosic degradation enzymes from extremophile organisms that show increased activity at extreme pH or temperature. These enzymes are inactive during normal plant growth to avoid detrimental effects and are activated under processing conditions such as elevated temperature or acidic pH. The nucleic acid sequences are optimized for expression in plants by removing problematic signals and codon optimization. Targeting sequences directing enzyme expression to specific cellular compartments improve the efficacy of biomass deconstruction upon activation.
Claims Coverage
The patent contains multiple independent claims focused on isolated polynucleotides, vectors, transformed host cells, plants, and methods of producing plants useful for biofuel production. The inventive features involve specific nucleic acid sequences, their encoded polypeptides, and associated targeting sequences.
Isolated polynucleotides encoding optimized cellulosic degradation enzymes
An isolated polynucleotide with at least 90% identity to sequences selected from SEQ ID NOs: 3-5, encoding a polypeptide at least 95% identical to SEQ ID NO: 13 or 14, which is an exogenous enzyme that hydrolyzes plant cell wall polysaccharides.
Vectors comprising polynucleotides with location-specific targeting sequences
A vector comprising the polynucleotide sequence together with a location-specific signal sequence that targets the encoded enzyme, such as an apoplast-targeting sequence with at least 90% identity to SEQ ID NO: 6.
Transformed host cells and plants containing the vectors
Host cells transformed with the described vectors and plants comprising such transformed host cells expressing the cellulosic degradation enzyme.
Methods for producing plants expressing extremophile-derived cellulosic degradation enzymes
A method of producing plants by introducing one or more exogenous nucleic acids encoding cellulosic degradation enzymes active at extreme pH or temperature, with sequences at least 90% identical to SEQ ID NOs: 3-5 and encoding polypeptides at least 95% identical to SEQ ID NO: 13 or 14, optionally using multiple nucleic acids encoding different enzymes, and optionally including location-specific signal sequences targeting enzyme localization to the apoplast.
Plants expressing exogenous cellulosic degradation enzymes from extremophiles
A biofuel production plant comprising one or more exogenous nucleic acids encoding cellulosic degradation enzymes from extremophile microorganisms, with nucleic acid sequences at least 90% identical to SEQ ID NOs: 3-5 and polypeptides at least 95% identical to SEQ ID NO: 13 or 14, optionally including location-specific targeting sequences for enzyme localization.
The claims cover isolated nucleic acids encoding optimized extremophile-derived enzymes for biomass hydrolysis, vectors and transformed cells/plants expressing these sequences, methods to produce such plants, and specific features including sequence identity thresholds and cellular targeting of enzymes, collectively providing comprehensive protection of the invention's components and methods.
Stated Advantages
Improved efficiency of biomass deconstruction due to expression of cellulosic degradation enzymes active under extreme conditions.
Reduced need for external enzyme reagents by producing enzymes within the biofuel feedstock plants.
Activation of enzymes only under processing conditions prevents detrimental effects during plant growth.
Optimization of nucleic acid sequences for plant expression enhances enzyme production and activity.
Targeting enzymes to specific cellular locations, such as the apoplast, enhances hydrolysis efficiency.
Documented Applications
Production of biofuels from lignocellulosic biomass by converting cellulosic and hemicellulosic polymers in plant cell walls into fermentable sugars using endogenous extremophile-derived enzymes expressed in transgenic plants.
Interested in licensing this patent?