Escherichia coli expressing chloroplast chaperones as a proxy to test heterologous Rubisco production in leaves

Rubisco is a fundamental enzyme in photosynthesis and therefore for life. Efforts to improve plant Rubisco performance have been hindered by the enzymes' complex chloroplast biogenesis requirements. New Synbio approaches, however, now allow the production of some plant Rubisco isoforms in Esche...

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Vydáno v:Journal of experimental botany Ročník 74; číslo 2; s. 664
Hlavní autoři: Buck, Sally, Rhodes, Tim, Gionfriddo, Matteo, Skinner, Tanya, Yuan, Ding, Birch, Rosemary, Kapralov, Maxim V, Whitney, Spencer M
Médium: Journal Article
Jazyk:angličtina
Vydáno: England 11.01.2023
Témata:
Carbon Dioxide - metabolism
Chloroplasts - metabolism
Escherichia coli - genetics
Escherichia coli - metabolism
Molecular Chaperones - metabolism
Nicotiana - metabolism
Photosynthesis
Plant Leaves - metabolism
Plants, Genetically Modified - metabolism
Ribulose-Bisphosphate Carboxylase - genetics
Ribulose-Bisphosphate Carboxylase - metabolism
Chloroplast transformation
synthetic biology
CO2 fixation
photosynthesis
Golden Gate cloning
ISSN:1460-2431, 1460-2431
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Shrnutí:Rubisco is a fundamental enzyme in photosynthesis and therefore for life. Efforts to improve plant Rubisco performance have been hindered by the enzymes' complex chloroplast biogenesis requirements. New Synbio approaches, however, now allow the production of some plant Rubisco isoforms in Escherichia coli. While this enhances opportunities for catalytic improvement, there remain limitations in the utility of the expression system. Here we generate, optimize, and test a robust Golden Gate cloning E. coli expression system incorporating the protein folding machinery of tobacco chloroplasts. By comparing the expression of different plant Rubiscos in both E. coli and plastome-transformed tobacco, we show that the E. coli expression system can accurately predict high level Rubisco production in chloroplasts but poorly forecasts the biogenesis potential of isoforms with impaired production in planta. We reveal that heterologous Rubisco production in E. coli and tobacco plastids poorly correlates with Rubisco large subunit phylogeny. Our findings highlight the need to fully understand the factors governing Rubisco biogenesis if we are to deliver an efficient, low-cost screening tool that can accurately emulate chloroplast expression.
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ISSN:1460-2431
1460-2431
DOI:10.1093/jxb/erac435