RT Journal Article
SR Electronic
T1 Resolving phylogenetic and biochemical barriers to functional expression of heterologous iron-sulphur cluster enzymes
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2021.02.02.429153
DO 10.1101/2021.02.02.429153
A1 Helena Shomar
A1 Pierre Simon Garcia
A1 Elena Fernández-Fueyo
A1 Francesca D’Angelo
A1 Martin Pelosse
A1 Rita Rebelo Manuel
A1 Ferhat Büke
A1 Siyi Liu
A1 Niels van den Broek
A1 Nicolas Duraffourg
A1 Carol de Ram
A1 Martin Pabst
A1 Simonetta Gribaldo
A1 Beatrice Py
A1 Sandrine Ollagnier de Choudens
A1 Gregory Bokinsky
A1 Frédéric Barras
YR 2021
UL http://biorxiv.org/content/early/2021/02/02/2021.02.02.429153.abstract
AB Many of the most promising applications of synthetic biology, including engineering of microbes for renewable chemical production, relies upon the ability of genetically-tractable hosts to express heterologous enzymes from foreign species. While countless methods for facilitating heterologous enzyme expression have been developed, comparable tools for facilitating heterologous enzyme activity are generally lacking. Such tools are needed to fully exploit the biosynthetic potential of the natural world. Here, using the model bacterium Escherichia coli, we investigate why iron-sulphur (Fe-S) enzymes are often inactive when heterologously expressed. By applying a simple growth complementation assay with collections of Fe-S enzyme orthologs from a wide range of prokaryotic diversity, we uncover a striking correlation between phylogenetic distance and probability of functional expression. Moreover, co-expression of a heterologous Fe-S biogenesis pathway increases the phylogenetic range of orthologs that can be functionally expressed. On the other hand, we find that heterologous Fe-S enzymes that require specific electron carrier proteins within their natural host are rarely functionally expressed unless their specific reducing partners are identified and co-expressed. We demonstrate in vitro that such selectivity in part derives from a need for low-potential electron donors. Our results clarify how phylogenetic distance and electron transfer biochemistry each separately impact functional heterologous expression and provide insight into how these barriers can be overcome for successful microbial engineering involving Fe-S enzymes.Competing Interest StatementThe authors have declared no competing interest.