RT Journal Article
SR Electronic
T1 Biosensor libraries harness large classes of binding domains for allosteric transcription regulators
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 193029
DO 10.1101/193029
A1 Juárez, Javier F.
A1 Lecube-Azpeitia, Begoña
A1 Brown, Stuart L.
A1 Church, George M.
YR 2017
UL http://biorxiv.org/content/early/2017/09/23/193029.abstract
AB Bacteria’s ability to specifically sense small molecules in their environment and trigger metabolic responses in accordance is an invaluable biotechnological resource. While many transcription factors (TFs) mediating these processes have been studied, only a handful has been leveraged for molecular biology applications. To expand this panel of biotechnologically important sensors here we present a strategy for the construction and testing of chimeric TF libraries, based on the fusion of highly soluble periplasmic binding proteins (PBPs) with DNA-binding domains (DBDs). We validated this strategy by constructing and functionally testing two unique sense-and-response regulators for benzoate, an environmentally and industrially relevant metabolite. This work will enable the development of tailored biosensors for synthetic regulatory circuits.3DThree-dimensionalApmRResistance to ampicillin/carbenicillinaTcAnhydrotetracyclineCmRResistance to chloramphenicolCRISPRClustered Regularly Interspaced Short Palindromic RepeatsCtC-terminalDBDDNA-Binding DomaindCas9Defective Cas9 protein, able to bind DNA and RNA but lacking nuclease activityeLCRenhanced LCRFACSFluorescence-Associated Cell SortingGGBPglucose-binding protein (E. coli)GmRResistance to gentamycinIPTGIsopropyl β-D-1-thiogalactopyranosideKmRResistance to kanamycinLCRLigase Chain ReactionLNKlinkernSPSBD that does Not contain its Signal PeptideNtn-terminalPBPPeriplasmic Binding ProteinPBSPhosphate Buffered SalineRBSRibosome Binding SiteSBDSubstrate-Binding DomainODOligomerization DomainPIDPercentage of IdentitySPSignal PeptideTALENsTranscription activator-like effector nucleaseTFTranscription factor