Cov khoom

Lub membrane topology ntawm colicin E1 channel sau tau kawm los ntawm fluorescence resonance zog hloov (FRET).Lub FRET koom nrog cov genetically encoded fluorescent amino acid (coumarin) ua tus pub thiab xaiv cov ntawv sau npe cysteine ​​residue tethered nrog DABMI (4-(dimethylamino) phenylazophenyl-4'-maleimide) ua tus txais FRET.Cov fluorescent coumarin residue tau muab tso rau hauv cov protein los ntawm ib qho orthogonal tRNA / aminoacyl-tRNA synthetase khub uas tso cai xaiv kev koom ua ke rau hauv ib qho chaw hauv colicin channel sau.Txhua qhov sib txawv tau muaj qhov nres (TAG) kev hloov pauv rau coumarin koom ua ke thiab cysteine ​​(TGT) hloov pauv rau DABMI txuas.Rau qhov kev ncua deb ntawm cov helices 1-6 tau txiav txim siab siv FRET tsom xam rau ob qho tib si soluble thiab membrane-bound xeev.Cov ntaub ntawv FRET tau pom tias muaj kev hloov pauv loj hauv qhov sib nrug deb ntawm cov helices 3-6 raws li lub koom haum membrane muab kev pom tshiab rau hauv daim nyias nyias-txheej txheem ntawm cov channel sau.Feem ntau, cov coumarin-DABMI FRET interhelical efficiencies txo qis raws li daim nyias nyias, ua raws li lub kaus qauv rau colicin channel.Tus qauv ntsuas rau lub xeev kaw ntawm cov channel sau tau tsim raws li cov ntaub ntawv FRET tam sim no thiab yav dhau los luam tawm.Tus qauv qhia txog lub voj voog ntawm helices 1-7 nyob rau hauv ib lub clockwise kev taw qhia los ntawm lub extracellular sab thiab membrane interfacial koom haum ntawm helices 1, 6, 7, thiab 10 nyob ib ncig ntawm lub central transmembrane hairpin tsim los ntawm helices 8 thiab 9.