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Title: Glycoside Hydrolases from a targeted Compost Metagenome, activity-screening and functional characterization

Abstract

Background: Metagenomics approaches provide access to environmental genetic diversity for biotechnology applications, enabling the discovery of new enzymes and pathways for numerous catalytic processes. Discovery of new glycoside hydrolases with improved biocatalytic properties for the efficient conversion of lignocellulosic material to biofuels is a critical challenge in the development of economically viable routes from biomass to fuels and chemicals.Results: Twenty-two putative ORFs (open reading frames) were identified from a switchgrass-adapted compost community based on sequence homology to related gene families. These ORFs were expressed in E. coli and assayed for predicted activities. Seven of the ORFs were demonstrated to encode active enzymes, encompassing five classes of hemicellulases. Four enzymes were over expressed in vivo, purified to homogeneity and subjected to detailed biochemical characterization. Their pH optima ranged between 5.5 - 7.5 and they exhibit moderate thermostability up to ~60-70°C.Conclusions: Seven active enzymes were identified from this set of ORFs comprising five different hemicellulose activities. These enzymes have been shown to have useful properties, such as moderate thermal stability and broad pH optima, and may serve as the starting points for future protein engineering towards the goal of developing efficient enzyme cocktails for biomass degradation under diverse process conditions.

Authors:
 [1];  [2];  [3];  [1];  [4];  [5]
  1. Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Sandia National Lab. (SNL-CA), Livermore, CA (United States)
  2. Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)
  3. Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States)
  4. Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Univ. of California, Berkeley, CA (United States)
  5. Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Sandia National Lab. (SNL-CA), Livermore, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)
Publication Date:
Research Org.:
Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Biological and Environmental Research (BER) (SC-23). Biological Systems Science Division
OSTI Identifier:
1511338
Grant/Contract Number:  
AC02-05CH11231
Resource Type:
Accepted Manuscript
Journal Name:
BMC Biotechnology (Online)
Additional Journal Information:
Journal Name: BMC Biotechnology (Online); Journal Volume: 12; Journal Issue: 1; Journal ID: ISSN 1472-6750
Publisher:
BioMed Central
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; Glycoside Hydrolase; Acetyl Esterase; Autoinduction Medium; Wheat Arabinoxylan; Endoxylanase Activity

Citation Formats

Dougherty, Michael J., D’haeseleer, Patrik, Hazen, Terry C., Simmons, Blake A., Adams, Paul D., and Hadi, Masood Z. Glycoside Hydrolases from a targeted Compost Metagenome, activity-screening and functional characterization. United States: N. p., 2012. Web. doi:10.1186/1472-6750-12-38.
Dougherty, Michael J., D’haeseleer, Patrik, Hazen, Terry C., Simmons, Blake A., Adams, Paul D., & Hadi, Masood Z. Glycoside Hydrolases from a targeted Compost Metagenome, activity-screening and functional characterization. United States. doi:10.1186/1472-6750-12-38.
Dougherty, Michael J., D’haeseleer, Patrik, Hazen, Terry C., Simmons, Blake A., Adams, Paul D., and Hadi, Masood Z. Tue . "Glycoside Hydrolases from a targeted Compost Metagenome, activity-screening and functional characterization". United States. doi:10.1186/1472-6750-12-38. https://www.osti.gov/servlets/purl/1511338.
@article{osti_1511338,
title = {Glycoside Hydrolases from a targeted Compost Metagenome, activity-screening and functional characterization},
author = {Dougherty, Michael J. and D’haeseleer, Patrik and Hazen, Terry C. and Simmons, Blake A. and Adams, Paul D. and Hadi, Masood Z.},
abstractNote = {Background: Metagenomics approaches provide access to environmental genetic diversity for biotechnology applications, enabling the discovery of new enzymes and pathways for numerous catalytic processes. Discovery of new glycoside hydrolases with improved biocatalytic properties for the efficient conversion of lignocellulosic material to biofuels is a critical challenge in the development of economically viable routes from biomass to fuels and chemicals.Results: Twenty-two putative ORFs (open reading frames) were identified from a switchgrass-adapted compost community based on sequence homology to related gene families. These ORFs were expressed in E. coli and assayed for predicted activities. Seven of the ORFs were demonstrated to encode active enzymes, encompassing five classes of hemicellulases. Four enzymes were over expressed in vivo, purified to homogeneity and subjected to detailed biochemical characterization. Their pH optima ranged between 5.5 - 7.5 and they exhibit moderate thermostability up to ~60-70°C.Conclusions: Seven active enzymes were identified from this set of ORFs comprising five different hemicellulose activities. These enzymes have been shown to have useful properties, such as moderate thermal stability and broad pH optima, and may serve as the starting points for future protein engineering towards the goal of developing efficient enzyme cocktails for biomass degradation under diverse process conditions.},
doi = {10.1186/1472-6750-12-38},
journal = {BMC Biotechnology (Online)},
number = 1,
volume = 12,
place = {United States},
year = {2012},
month = {7}
}

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Works referenced in this record:

Glycoside Hydrolase Activities of Thermophilic Bacterial Consortia Adapted to Switchgrass
journal, July 2011

  • Gladden, John M.; Allgaier, Martin; Miller, Christopher S.
  • Applied and Environmental Microbiology, Vol. 77, Issue 16, p. 5804-5812
  • DOI: 10.1128/AEM.00032-11

Protein engineering in designing tailored enzymes and microorganisms for biofuels production
journal, August 2009


Xylanases, xylanase families and extremophilic xylanases
journal, January 2005


Protein production by auto-induction in high-density shaking cultures
journal, May 2005