Journal article
Functional mining of transporters using synthetic selections
Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark1
Research Groups, Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark2
Bacterial Synthetic Biology, Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark3
Synthetic Biology Tools for Yeast, Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark4
Biosyntia ApS5
iLoop, Translational Management, Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark6
Only 25% of bacterial membrane transporters have functional annotation owing to the difficulty of experimental study and of accurate prediction of their function. Here we report a sequence-independent method for high-throughput mining of novel transporters. The method is based on ligand-responsive biosensor systems that enable selective growth of cells only if they encode a ligand-specific importer.
We developed such a synthetic selection system for thiamine pyrophosphate and mined soil and gut metagenomes for thiamine-uptake functions. We identified several members of a novel class of thiamine transporters, PnuT, which is widely distributed across multiple bacterial phyla. We demonstrate that with modular replacement of the biosensor, we could expand our method to xanthine and identify xanthine permeases from gut and soil metagenomes.
Our results demonstrate how synthetic-biology approaches can effectively be deployed to functionally mine metagenomes and elucidate sequence–function relationships of small-molecule transport systems in bacteria.
Language: | English |
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Publisher: | Nature Publishing Group US |
Year: | 2016 |
Pages: | 1015-1022 |
ISSN: | 15524469 and 15524450 |
Types: | Journal article |
DOI: | 10.1038/nchembio.2189 |
ORCIDs: | Genee, Hans Jasper , Petersen, Søren Dalsgård , Siedler, Solvej , Kristensen, Mette and Sommer, Morten Otto Alexander |