Download - ))sors - Ghent University Academic Bibliography · Introduction "If you cannot measure it, you cannot improve it."gSgKelvinIgB@gAOgMeasuremenggggmetaboggessentialggmetabogengineeringgg

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Page 1: ))sors - Ghent University Academic Bibliography · Introduction "If you cannot measure it, you cannot improve it."gSgKelvinIgB@gAOgMeasuremenggggmetaboggessentialggmetabogengineeringgg

Introduction

"If you cannot measure it, you cannot improve it."g SLordgKelvinIgB@thgcenturyAOgMeasurementgofgkeygpathwaygmetabolitesg isgessentialg ingmetabolicgengineeringg togensuregthegdevelopmentgofgefficientgandgoptimalgmicrobialgproductiongstrainsOg/urrentgmethodsgforganalysisgofgintracellulargmetabolitesgaregcumbersomegandgdemandgexgvivoIgdestructivegtechniquesgsuchgasgHPL/IgG/gandgL/&MSOg:sgangelegantgsolutionIg transcriptionalgbiosensorsIgconsistinggofg transcriptionalgregulatorygcircuitsgfoundg ingnatureIg cangbegusedOgThesegbiosensorsgenableg ing vivoIg non&destructiveIg proportionalIg high&throughputgandgevengsimultaneousgmeasurementgofg specificgmetabolitesgofginterestO

Thegauthorsgwishgtogthankgthegx:gentschapgInnoverengGgOndernemenxgingFlandersgS:IOAgforgfinancialgsupportgingthegframeworkgofgthegPhODOggrantgofgWrechtgDegPaepeOgThisgresearchgisgalsogsupportedgbygthegMultidisciplinarygResearchgPartnershipgGhentgWio&Economygandgtheg/ondexgprojectgSGODERBOBENAOg

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Acknowledgments Contact

Tailor-made)Transcriptional)BiosensorsWrechtgDegPaepeBIgJogMaertensBIgWartelgVanholmeRgandgMarjangDegMeyB

BMEMOg&gMetabolicgEngineeringgGroupg&gLaboratorygofgIndustrialgWiotechnologygandgWiocatalysisg&gGhentgUniversityg&gWelgiumRVIWg&gVlaamsgInstituutgvoorgWiotechnologieg&gDepartmentgofgPlantgSystemsgWiologyg&gWegliumg

Tailor-made)characteristic)response)curve

Transcription)factor Transcription)factor)binding)sites

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Biosensor)copy)number

Chimeric)transcriptional)regulation)pairs Chimeric)transcription)factors

Current)results

Theg ratiog ofg transcriptiong factorg expressiong versusg theg copyg numberg ofg theg bindingg sitesg andg thegintracellulargconcentrationgofgthegspecificgligandgplaysgagkeygrolegingdefininggthegbiosensorgresponsegcurveOgWygalteringgthegpromotergandgRWSgstrengthgofgthegtranscriptiongfactorgoperonIgthegresponsegcurvegcangbegalteredO

Engineeringgthegnativegbiosensorgpartsgenablesgthegdevelopmentgofguniquegbiosensorsgwithgcharacteristicgresponsegcurvesgcustomizedgtogthegresearcher’sgneedsOgTranscriptionalg biosensorg architectureg cang beg dividedg intog threeg basicg levelsg ofg controlUg (1)g theg transcriptiong factorg operonIg (2)g theg reporterg operong withg thegtranscriptiongfactorgbindinggsitesgandg(3)gthegSplasmidAgcopygnumbergofgthegcompletegbiosensorOgggg

Engineeringgtranscriptiongfactorsgenablesgthegdevelopmentgofgbiosensorsgwithguniquegmolecule&specificitiesgcustomizedgtogthegresearcher’sgneedsOgWithingthegsamegtranscriptiongfactorgfamilyIghighlygconservedgDN:&gandgligand&bindinggdomainsgaregdistinguishableOg:dditionallyIgthegcorrespondinggtranscriptiongfactorgbindinggsitesgalsog showg highg conservationOg ThereforeIg completeg transcriptiong factorsg org eveng specificg ligand&bindingg proteing domainsg areg interchangedg withg ag Spartg ofAg thegtranscriptiongfactorgofgagcharaterizedgbiosensorgtogcreategchimericgbiosensorsgwithgothergligandgspecificitiesgbutgpredictablegresponsegcurvesO

FluorescentproteinPromoterTranscriptionfactorPromoter

ThegSplasmidAgcopygnumbergofgthegbiosensorgdirectlygeffectsgthegtotalgexpressiongofgbothgtranscriptiongfactorgandgfluorescentgreportergproteingandgthegcopygnumbergofgthegtranscriptiongfactorgbindinggsitesOg/onsequentlyIgthegbiosensorgcopygnumbergco&definesgthegbiosensorgresponsegcurveOgggg

Dueg tog theghighgconservationgofgDN:&bindinggsitesgacrossgdifferentgspeciesIgcombininggag ligandgspecificgtranscriptiongfactorgfromgonegspeciesgwithgDN:&bindinggsitesgfromgagcharacterizedgbiosensorIgcangcreategagfunctionalg chimericg bioesensorg withg differentg ligandg specificityOg Theseg chimericg biosensorsg onlyg differg ingtheirgtranscriptiongfactorIgthusgenhancinggconstructiongefficiencygandgresponsegcurvegpredictabilityO

Transcriptiong factorsg withg conservedg andg distinctg DN:&g andg ligand&bindingg proteing domainsg andg withgdifferentgligandgspecificitiesgaregusedgtogconstructgchimericgtranscriptiongfactorsOgThereforeIgagcharacterizedgbiosensorg withg ag definedg responseg curveg isg usedg asg ag chassisg forg constructingg ag predictableg chimericgbiosensorgwithgagchimericgtranscriptiongfactorgthatgcontainsgthegsubstrategbindinggdomaingwithgthegdesiredgspecificityOg

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