Combinatorial design of chemical-dependent protein switches for controlling intracellular electron transfer

Bingyan Wu, Joshua T. Atkinson, Dimithree Kahanda, George N. Bennett, Jonathan J. Silberg

Research output: Contribution to journalArticlepeer-review

8 Scopus citations

Abstract

One challenge with controlling electron flow in cells is the lack of biomolecules that directly couple environmental sensing to electron transfer efficiency. To overcome this component limitation, we randomly inserted the ligand binding domain (LBD) from the estrogen receptor (ER) into a 2Fe-2S ferredoxin (Fd) and used a bacterial selection to identify protein variants that support electron transfer in cells. Mapping LBD insertion sites onto structure revealed that Fd tolerates domain insertion adjacent to or within the tetracysteine motif that coordinates the 2Fe-2S metallocluster. With both protein designs, cellular electron transfer (ET) was enhanced by the ER antagonist 4-hydroxytamoxifen, albeit to different extents. One variant arising from ER-LBD insertion within the tetracysteine motif acquired an oxygen-tolerant 2Fe-2S cluster, suggesting that ET is regulated through post-translational ligand binding. These metalloprotein switches are expected to be useful for achieving fast regulation of ET in engineered metabolic pathways and between electroactive bacteria and conductive materials.

Original languageEnglish (US)
Article numbere16796
JournalAIChE Journal
Volume66
Issue number3
DOIs
StatePublished - Mar 1 2020
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Biotechnology
  • Environmental Engineering
  • General Chemical Engineering

Keywords

  • allostery
  • domain insertion
  • electron transfer
  • ferredoxin
  • growth selection

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