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Recombination of 2Fe-2S Ferredoxins Reveals Differences in the Inheritance of Thermostability and Midpoint Potential. | LitMetric

AI Article Synopsis

  • * Researchers created chimeric ferredoxins (Fd) by mixing genes from cyanobacterial and cyanomyophage sources, resulting in proteins with varying amino acid compositions, thermostabilities, and modified electrochemical properties.
  • * The study showed that these synthetic Fds can support electron transfer in metabolic pathways, revealing potential for tailored enzyme development and insights into the inheritance of traits like stability and activity.

Article Abstract

Recombination can be used in the laboratory to overcome component limitations in synthetic biology by creating enzymes that exhibit distinct activities and stabilities from native proteins. To investigate how recombination affects the properties of an oxidoreductase that transfers electrons in cells, we created ferredoxin (Fd) chimeras by recombining distantly related cyanobacterial and cyanomyophage Fds (53% identity) that present similar midpoint potentials but distinct thermostabilities. Fd chimeras having a wide range of amino acid substitutions retained the ability to coordinate an iron-sulfur cluster, although their thermostabilities varied with the fraction of residues inherited from each parent. The midpoint potentials of chimeric Fds also varied. However, all of the synthetic Fds exhibited midpoint potentials outside of the parental protein range. Each of the chimeric Fds could also support electron transfer between Fd-NADP reductase and sulfite reductase in , although the chimeric Fds varied in the expression required for similar levels of cellular electron transfer. These results show how Fds can be diversified through recombination and reveal differences in the inheritance of thermostability and electrochemical properties. Furthermore, they illustrate how electron transfer efficiencies of chimeric Fds can be rapidly evaluated using a synthetic metabolic pathway.

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Source
http://dx.doi.org/10.1021/acssynbio.0c00303DOI Listing

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