Method for reducing viscosity in saccharification process

a technology of viscosity reduction and saccharification process, which is applied in the direction of hydrolases, biochemical apparatus and processes, enzymes, etc., can solve the problems of slow ethanol conversion research, inability to achieve efficient degradation of whole cellulase, and inability to achieve natural-occurring whole cellulase, etc., to achieve the yield of desirable fermentable sugars, reduce the viscosity of the mixture, and improve the yield of saccharification

Inactive Publication Date: 2014-05-15
DANISCO US INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0018]The present disclosure is based, in part, on the surprising discovery that inclusion of a certain endoglucanase enzyme (e.g., a polypeptide having glycosyl hydrolase family 61 (“GH61”)/endoglucanase activity, such as the T. reesei endoglucanase (“Eg4”)) in a b...

Problems solved by technology

These early processes were primarily directed to acid hydrolysis, which were complex in engineering and design, and were typically sensitive to small variations in the processes, such as to temperature, pressure and/or acid concentrations.
The abundant supply of petroleum in the period from World War II through the early 1970s slowed ethanol conversion research.
Uncertainty remains with these processes, mainly on their ability to be scaled up for commercialization and on the efficiency of ethanol production.
Organisms are often restrictive with regard to which sugars they use, and this dictates which sugars are best to produce during conversion.
A whole cellulase, especially one that is naturally occurring, is, however, not necessarily capable of achieving efficient degradation because it may not include all the components/activities required for this efficiency, for example, activities from each of the CBH, EG and BG c...

Method used

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  • Method for reducing viscosity in saccharification process
  • Method for reducing viscosity in saccharification process
  • Method for reducing viscosity in saccharification process

Examples

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example 1

Assays / Methods

[0360]The following assays / methods were generally used in the Examples described below. Any deviations from the protocols provided below are indicated in specific Examples.

[0361]A. Pretreatment of Biomass Substrates

[0362]Corncob, corn stover and switch grass were pretreated prior to enzymatic hydrolysis according to the methods and processing ranges described in International Patent Publication WO06110901A (unless otherwise noted). These references for pretreatment are also included in the disclosures of US Patent Application Publications 20070031918-A1, 20070031919-A1, 20070031953-A1, and / or 20070037259-A1.

[0363]Ammonia fiber explosion treated (AFEX) corn stover was obtained from Michigan Biotechnology Institute International (MBI). The composition of the corn stover was determined by MBI (Teymouri, F et al. Applied Biochemistry and Biotechnology, 2004, 113:951-963) using the National Renewable Energy Laboratory (NREL) procedure, NREL LAP-002. NREL procedures are avai...

example 2

Construction of an Integrated Expression Strain of Trichoderma reesei

[0386]An integrated expression strain of Trichoderma reesei was constructed that co-expressed five genes: T. reesei β-glucosidase gene bgl1, T. reesei endoxylanase gene xyn3, F. verticillioides β-xylosidase gene fv3A, F. verticillioides β-xylosidase gene fv43D, and F. verticillioides α-arabinofuranosidase gene fv51A.

[0387]The construction of the expression cassettes for these different genes and the transformation of T. reesei are described below.

[0388]A. Construction of the β-Glucosidase Expression Vector

[0389]The N-terminal portion of the native T. reesei β-glucosidase gene bgl1 was codon optimized by DNA 2.0 (Menlo Park, USA). This synthesized portion comprised of the first 447 bases of the coding region. This fragment was PCR amplified using primers SK943 and SK941. The remaining region of the native bgl1 gene was PCR amplified from a genomic DNA sample extracted from T. reesei strain RL-P37 (Sheir-Neiss, G et...

example 3

Construction of T. reesei Strains

[0414]A. Construction of and Screening for T. Reesei Strain H3A / EG4#27

[0415]An expression cassette containing the T. reesei egl1 (also termed “Cel 7B”) promoter, T. reesei eg4 (also termed “TrEG4”, or “Cel 61A”) open reading frame, and cbh1 (Cel 7A) terminator sequence (FIG. 12A) from Trichoderma reesei, and sucA selectable marker (see, Boddy et al., Curr. Genet. 1993, 24:60-66) from Aspergillus niger was cloned into pCR Blunt II TOPO (Invitrogen) (FIG. 12B).

[0416]The expression cassette Pegl1-eg4-sucA was amplified by PCR with the primers:

(SEQ ID NO: 146)SK1298: 5′-GTAGTTATGCGCATGCTAGAC-3′(SEQ ID NO: 147)214: 5′-CCGGCTCAGTATCAACCACTAAGCACAT-3′

[0417]Pfu Ultra II (Stratagene) was used as the polymerase for the PCR reaction. The products of the PCR reaction were purified with the QIAquick PCR purification kit (Qiagen) as per the manufacturer's protocol. The products of the PCR reaction were then concentrated using a speed vac to 1-3 μg / μL. The T. reese...

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Abstract

The present invention relates to compositions that can be used in hydrolyzing biomass such as compositions comprising a polypeptide having glycosyl hydrolase family 61/endoglucanase activity, methods for hydrolyzing biomass material, and methods for reducing viscosity of biomass mixture using a composition comprising a polypeptide having glycosyl hydrolase family 61/endoglucanase activity.

Description

CROSS REFERENCE TO RELATED APPLICATIONS[0001]This application claims the benefit of U.S. Provisional Application No. 61 / 453,923, filed Mar. 17, 2011, which is hereby incorporated by reference in its entirety.FIELD OF THE INVENTION[0002]The present invention relates to compositions useful for hydrolyzing biomass, methods of using such compositions to hydrolyze biomass materials, and methods for reducing viscosity of biomass saccharification mixtures.BACKGROUND OF THE INVENTION[0003]Bioconversion of renewable lignocellulosic biomass to a fermentable sugar that is subsequently fermented to produce alcohol (e.g., ethanol) as an alternative to liquid fuels has attracted the intensive attention of researchers since the 1970s, when the oil crisis occurred (Bungay, H. R., “Energy: the biomass options”. NY: Wiley; 1981; Olsson L, Hahn-Hagerdal B. Enzyme Microb Technol 1996, 18:312-31; Zaldivar, J et al., Appl Microbiol Biotechnol 2001, 56: 17-34; Galbe, M et al., Appl Microbiol Biotechnol 20...

Claims

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Application Information

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IPC IPC(8): C12P19/02C12P19/14
CPCC12P19/14C12P19/02C12N9/2437C12Y302/01004C12N9/24C12N15/52
Inventor MITCHINSON, COLINLI, MIANKELEMEN, BRADLEY R.LANTZ, SUZANNE E.WING, KEITH D.HITZ, WILLIAM D.
Owner DANISCO US INC
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