Structural investigation of the transmembrane C domain of the mannitol permease from Escherichia coli using 5-FTrp fluorescence spectroscopy

Autor: Jaap Broos, Fabrizia Fusetti, Milena Opacic, Ben H. Hesp, Bauke W. Dijkstra
Přispěvatelé: Enzymology, X-ray Crystallography
Rok vydání: 2011
Předmět:
PTS
Monosaccharide Transport Proteins
Stereochemistry
Dimer
Amino Acid Motifs
Biophysics
Biological Transport
Active
CYTOPLASMIC B-DOMAIN
SUBSTRATE-BINDING
Plasma protein binding
medicine.disease_cause
Biochemistry
chemistry.chemical_compound
medicine
Escherichia coli
Inner membrane
ENZYME-IIMTL
Mannitol
Binding site
PHOSPHORYLATION SITE MUTANTS
TRANSPORT-SYSTEM
Phosphorylation
Fluorotryptophan
Phosphoenolpyruvate Sugar Phosphotransferase System
Protein Structure
Quaternary
Time-resolved fluorescence spectroscopy
biology
Chemistry
MEMBRANE-PROTEINS
ACTIVE-SITE
Escherichia coli Proteins
Tryptophan
Active site
Iodide quenching
Cell Biology
Periplasmic space
Transmembrane protein
Protein Structure
Tertiary
Spectrometry
Fluorescence
DEPENDENT PHOSPHOTRANSFERASE SYSTEM
PHOSPHOENOLPYRUVATE
Membrane protein
biology.protein
Anisotropy
psychological phenomena and processes
Protein Binding
Zdroj: Biochimica et Biophysica Acta-Biomembranes, 1818(3), 861-868. Elsevier
ISSN: 0006-3002
0005-2736
Popis: The mannitol transporter EIImtl from Escherichia coli is responsible for the uptake of mannitol over the inner membrane and its concomitant phosphorylation. EIImtl is functional as a dimer and its membrane-embedded C domain, IICmtl, harbors one high affinity mannitol binding site. To characterize this domain in more detail the microenvironments of thirteen residue positions were explored by 5-fluorotryptophan (5-FTrp) fluorescence spectroscopy. Because of the simpler photophysics of 5-FTrp compared to Trp, one can distinguish between the two 5-FTrp probes present in dimeric IICmtl. At many labeled positions, the microenvironment of the 5-FTrps in the two protomers differs. Spectroscopic properties of three mutants labeled at positions 198. 251, and 260 show that two conserved motifs (Asn194-His195 and Gly254-Ile255-His256-Glu257) are located in well-structured parts of IICmtl. Mannitol binding has a large impact on the structure around position 198. while only minor changes are induced at positions 251 and 260. Phosphorylation of the cytoplasmic B domain of EIImtl is sensed by 5-FTrp at positions 30, 42, 251 and 260. We conclude that many parts of the IICmtl structure are involved in the sugar translocation. The structure of EIImtl, as investigated in this work. differs from the recently solved structure of a IIC protein transporting diacetylchitobiose, ChbC, and also belonging to the glucose superfamily of EII sugar transporters. In EIImtl, the sugar binding site is more close to the periplasmic face and the structure of the 2 protomers in the dimer is different, while both protomers in the ChbC dimer are essentially the same. (C) 2011 Elsevier B.V. All rights reserved.
Databáze: OpenAIRE
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