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Title: Functional metagenomic selection of ribulose 1, 5-bisphosphate carboxylase/oxygenase from uncultivated bacteria

Abstract

Ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) is a critical yet severely inefficient enzyme that catalyses the fixation of virtually all of the carbon found on Earth. Here, we report a functional metagenomic selection that recovers physiologically active RubisCO molecules directly from uncultivated and largely unknown members of natural microbial communities. Selection is based on CO 2-dependent growth in a host strain capable of expressing environmental deoxyribonucleic acid (DNA), precluding the need for pure cultures or screening of recombinant clones for enzymatic activity. Seventeen functional RubisCO-encoded sequences were selected using DNA extracted from soil and river autotrophic enrichments, a photosynthetic biofilm and a subsurface groundwater aquifer. Notably, three related form II RubisCOs were recovered which share high sequence similarity with metagenomic scaffolds from uncultivated members of the Gallionellaceae family. One of the Gallionellaceae Rubis COs was purified and shown to possess CO 2/O 2 specificity typical of form II enzymes. X-ray crystallography determined that this enzyme is a hexamer, only the second form II multimer ever solved and the first RubisCO structure obtained from an uncultivated bacterium. Functional metagenomic selection leverages natural biological diversity and billions of years of evolution inherent in environmental communities, providing a new window into the discovery of COmore » 2-fixing enzymes not previously characterized.« less

Authors:
 [1];  [1];  [1];  [2];  [3];  [4];  [5];  [6];  [6];  [4];  [1];  [1]
  1. The Ohio State Univ., Columbus, OH (United States). Dept. of Microbiology
  2. Botanical Research Inst. of Texas, Fort Worth,TX (United States)
  3. Univ. of Sao Paulo, Sao Paulo (Brazil). Dept. of Genetics, ESALQ
  4. Univ. of California, Berkeley, CA (United States). Dept. of Earth and Planetary Science
  5. Univ. of California, Los Angeles, CA (United States). Protein Expression Technology Center, UCLA-DOE Inst.
  6. U.S. Geological Survey, Menlo Park, 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); National Institutes of Health (NIH)
OSTI Identifier:
1471013
Grant/Contract Number:  
AC02-05CH11231; FC02‐02ER63421
Resource Type:
Journal Article: Accepted Manuscript
Journal Name:
Environmental Microbiology
Additional Journal Information:
Journal Volume: 18; Journal Issue: 4; Journal ID: ISSN 1462-2912
Publisher:
Wiley
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES

Citation Formats

Varaljay, Vanessa A., Satagopan, Sriram, North, Justin A., Witte, Brian, Dourado, Manuella N., Anantharaman, Karthik, Arbing, Mark A., McCann, Shelley Hoeft, Oremland, Ronald S., Banfield, Jillian F., Wrighton, Kelly C., and Tabita, F. Robert. Functional metagenomic selection of ribulose 1, 5-bisphosphate carboxylase/oxygenase from uncultivated bacteria. United States: N. p., 2015. Web. doi:10.1111/1462-2920.13138.
Varaljay, Vanessa A., Satagopan, Sriram, North, Justin A., Witte, Brian, Dourado, Manuella N., Anantharaman, Karthik, Arbing, Mark A., McCann, Shelley Hoeft, Oremland, Ronald S., Banfield, Jillian F., Wrighton, Kelly C., & Tabita, F. Robert. Functional metagenomic selection of ribulose 1, 5-bisphosphate carboxylase/oxygenase from uncultivated bacteria. United States. doi:10.1111/1462-2920.13138.
Varaljay, Vanessa A., Satagopan, Sriram, North, Justin A., Witte, Brian, Dourado, Manuella N., Anantharaman, Karthik, Arbing, Mark A., McCann, Shelley Hoeft, Oremland, Ronald S., Banfield, Jillian F., Wrighton, Kelly C., and Tabita, F. Robert. Mon . "Functional metagenomic selection of ribulose 1, 5-bisphosphate carboxylase/oxygenase from uncultivated bacteria". United States. doi:10.1111/1462-2920.13138. https://www.osti.gov/servlets/purl/1471013.
@article{osti_1471013,
title = {Functional metagenomic selection of ribulose 1, 5-bisphosphate carboxylase/oxygenase from uncultivated bacteria},
author = {Varaljay, Vanessa A. and Satagopan, Sriram and North, Justin A. and Witte, Brian and Dourado, Manuella N. and Anantharaman, Karthik and Arbing, Mark A. and McCann, Shelley Hoeft and Oremland, Ronald S. and Banfield, Jillian F. and Wrighton, Kelly C. and Tabita, F. Robert},
abstractNote = {Ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) is a critical yet severely inefficient enzyme that catalyses the fixation of virtually all of the carbon found on Earth. Here, we report a functional metagenomic selection that recovers physiologically active RubisCO molecules directly from uncultivated and largely unknown members of natural microbial communities. Selection is based on CO2-dependent growth in a host strain capable of expressing environmental deoxyribonucleic acid (DNA), precluding the need for pure cultures or screening of recombinant clones for enzymatic activity. Seventeen functional RubisCO-encoded sequences were selected using DNA extracted from soil and river autotrophic enrichments, a photosynthetic biofilm and a subsurface groundwater aquifer. Notably, three related form II RubisCOs were recovered which share high sequence similarity with metagenomic scaffolds from uncultivated members of the Gallionellaceae family. One of the Gallionellaceae Rubis COs was purified and shown to possess CO2/O2 specificity typical of form II enzymes. X-ray crystallography determined that this enzyme is a hexamer, only the second form II multimer ever solved and the first RubisCO structure obtained from an uncultivated bacterium. Functional metagenomic selection leverages natural biological diversity and billions of years of evolution inherent in environmental communities, providing a new window into the discovery of CO2-fixing enzymes not previously characterized.},
doi = {10.1111/1462-2920.13138},
journal = {Environmental Microbiology},
issn = {1462-2912},
number = 4,
volume = 18,
place = {United States},
year = {2015},
month = {11}
}

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