Spin labeling analysis of structure and dynamics of the Na+/proline transporter of Escherichia coli

DC FieldValueLanguage
dc.contributor.authorWegener, C
dc.contributor.authorTebbe, S
dc.contributor.authorSteinhoff, HJ
dc.contributor.authorJung, HR
dc.date.accessioned2021-12-23T16:07:50Z-
dc.date.available2021-12-23T16:07:50Z-
dc.date.issued2000
dc.identifier.issn00062960
dc.identifier.urihttps://osnascholar.ub.uni-osnabrueck.de/handle/unios/8088-
dc.description.abstractWith respect to the functional importance attributed to the N-terminal part of the Na+/proline transporter of Escherichia coli (PutP), we report here on the structural arrangement and functional dynamics of transmembrane domains (TMs) II and III and the adjoining loop regions. Information on membrane topography was obtained by analyzing the residual mobility of site-specifically-attached nitroxide spin label and by determination of collision frequencies of the nitroxide with oxygen and a polar metal ion complex using electron paramagnetic resonance (EPR) spectroscopy. The studies suggest that amino acids Phe45, Ser50, Ser54, Trp59, and Met62 are part of TM II while Gly39 and Arg40 are located at a membrane-water interface probably forming the cytoplasmic cap of the TM. Also Ala67 and Glu75 are at a membrane-water interface, suggesting a location close to the periplasmic ends of TMs II and III, respectively. Ser71 between these residues is clearly in a water-exposed loop (periplasmic loop 3). Spin labels attached to positions 80, 86, and 91 show EPR properties typical for a TM location (TM III). Leu97 may be part of a structured loop region while Ala107 is clearly located in a water-exposed loop (cytoplasmic loop 4). Finally, spin labels attached to the positions of Asp33 and Leu37 are clearly on the surface of the transporter and are directed into an apolar environment. These findings strongly support the recently proposed 13-helix model of PutP [Jung, H., Rubenhagen, R., Tebbe, S., Leifker, K,, Tholema, N., Quick, M., and Schmid, R. (1998) J. Biol. Chern. 273, 26400-26407] and suggest that TMs II and III of the transporter are formed by amino acids Ser41 to Gly66 and Ser76 to Gly95, respectively. In addition to the topology analysis, it is shown that binding of Na+ and/or proline to the transporter alters the mobility of the nitroxide group at the positions of Leu37 and Phe45. From these findings, it is concluded that binding of the ligands induces conformational alterations of PutP that involve at least parts of TM II and the preceding cytoplasmic loop.
dc.language.isoen
dc.publisherAMER CHEMICAL SOC
dc.relation.ispartofBIOCHEMISTRY
dc.subjectBACTERIORHODOPSIN MUTANTS
dc.subjectBiochemistry & Molecular Biology
dc.subjectCOTRANSPORTER FAMILY
dc.subjectELECTRON-PARAMAGNETIC-RESONANCE
dc.subjectINTERSPIN DISTANCES
dc.subjectLACTOSE PERMEASE
dc.subjectLIGHT-DEPENDENT CHANGES
dc.subjectLINKING IN-SITU
dc.subjectMEMBRANE-PROTEINS
dc.subjectPROLINE UPTAKE
dc.subjectSIDE-CHAINS
dc.titleSpin labeling analysis of structure and dynamics of the Na+/proline transporter of Escherichia coli
dc.typejournal article
dc.identifier.doi10.1021/bi992442x
dc.identifier.isiISI:000086737700030
dc.description.volume39
dc.description.issue16
dc.description.startpage4831
dc.description.endpage4837
dc.contributor.orcid0000-0002-5888-0157
dc.contributor.orcid0000-0002-5450-3063
dc.contributor.researcheridH-3791-2014
dc.contributor.researcheridK-3790-2014
dc.publisher.place1155 16TH ST, NW, WASHINGTON, DC 20036 USA
dcterms.isPartOf.abbreviationBiochemistry
crisitem.author.deptFB 04 - Physik-
crisitem.author.deptidfb04-
crisitem.author.parentorgUniversität Osnabrück-
crisitem.author.netidStHe633-
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