TY - JOUR
T1 - Functional mining of transporters using synthetic selections
AU - Genee, Hans Jasper
AU - Bali, Anne Pihl
AU - Petersen, Søren Dalsgård
AU - Siedler, Solvej
AU - Bonde, Mads
AU - Gronenberg, Luisa S.
AU - Kristensen, Mette
AU - Harrison, Scott James
AU - Sommer, Morten Otto Alexander
PY - 2016
Y1 - 2016
N2 - 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.
AB - 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.
U2 - 10.1038/nchembio.2189
DO - 10.1038/nchembio.2189
M3 - Journal article
C2 - 27694800
SN - 1552-4450
JO - Nature Chemical Biology
JF - Nature Chemical Biology
ER -