Directed -in vitro- Evolution of precambrian and extant rubiscos

Bernardo J. Gomez-Fernandez, Eva Garcia-Ruiz, Javier Martin-Diaz, Patricia Gomez De Santos, Paloma Santos-Moriano, Francisco J. Plou, Antonio Ballesteros, Monica Garcia, Marisa Rodriguez, Valeria A. Risso, Jose M. Sanchez-Ruiz, Spencer M. Whitney, Miguel Alcalde*

*Corresponding author for this work

    Research output: Contribution to journalArticlepeer-review

    20 Citations (Scopus)

    Abstract

    Rubisco is an ancient, catalytically conserved yet slow enzyme, which plays a central role in the biosphere's carbon cycle. The design of Rubiscos to increase agricultural productivity has hitherto relied on the use of in vivo selection systems, precluding the exploration of biochemical traits that are not wired to cell survival. We present a directed -in vitro- evolution platform that extracts the enzyme from its biological context to provide a new avenue for Rubisco engineering. Precambrian and extant form II Rubiscos were subjected to an ensemble of directed evolution strategies aimed at improving thermostability. The most recent ancestor of proteobacteria -dating back 2.4 billion years- was uniquely tolerant to mutagenic loading. Adaptive evolution, focused evolution and genetic drift revealed a panel of thermostable mutants, some deviating from the characteristic trade-offs in CO2-fixing speed and specificity. Our findings provide a novel approach for identifying Rubisco variants with improved catalytic evolution potential.

    Original languageEnglish
    Article number5532
    JournalScientific Reports
    Volume8
    Issue number1
    DOIs
    Publication statusPublished - 1 Dec 2018

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