Mapping of the Dimer Interface of the Escherichia coli Mannitol Permease by Cysteine Cross-linking
| Autor: | Jaap Broos, Gea K. Schuurman-Wolters, Joyce Wind, George T. Robillard, Bart A. van Montfort, Bert Poolman |
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| Přispěvatelé: | X-ray Structuurbepaling, Faculty of Science and Engineering, Enzymologie, Groningen Biomolecular Sciences and Biotechnology |
| Rok vydání: | 2002 |
| Předmět: |
Monosaccharide Transport Proteins
Dimer Molecular Sequence Data ANGSTROM RESOLUTION Biochemistry chemistry.chemical_compound Escherichia coli ENZYME-IIMTL Amino Acid Sequence Cysteine Phosphoenolpyruvate Sugar Phosphotransferase System PHOSPHORYLATION Site-directed mutagenesis GeneralLiterature_REFERENCE(e.g. dictionaries encyclopedias glossaries) Molecular Biology Peptide sequence TRANSPORT PROTEIN Escherichia coli Proteins Cell Biology PROJECTION STRUCTURE TRANSMEMBRANE DOMAIN Transport protein Transmembrane domain Crystallography Monomer Membrane chemistry DEPENDENT PHOSPHOTRANSFERASE SYSTEM SITE-SPECIFIC MUTAGENESIS PHOSPHOENOLPYRUVATE ComputingMethodologies_DOCUMENTANDTEXTPROCESSING Mutagenesis Site-Directed MEMBRANE Dimerization |
| Zdroj: | The Journal of Biological Chemistry, 277(17), 14717-14723. AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC |
| ISSN: | 0021-9258 |
| DOI: | 10.1074/jbc.m201533200 |
| Popis: | A cysteine cross-linking approach was used to identify residues at the dimer interface of the Escherichia coli mannitol permease. This transport protein comprises two cytoplasmic domains and one membrane-embedded C domain per monomer, of which the latter provides the dimer contacts. A series of single-cysteine His-tagged C domains present in the native membrane were subjected to Cu(II)-(1,10-phenanthroline)(3)-catalyzed disulfide formation or cysteine cross-linking with dimaleimides of different length. The engineered cysteines were at the borders of the predicted membrane-spanning alpha-helices. Two residues were found to be located in close proximity of each other and capable of forming a disulfide, while four other locations formed cross-links with the longer dimaleimides. Solubilization of the membranes did only influence the cross-linking behavior at one position (Cys(73)). Mannitol binding only effected the cross-linking of a cysteine at the border of the third transmembrane helix (Cys(134)), indicating that substrate binding does not lead to large rearrangements in the helix packing or to dissociation of the dimer. Upon mannitol binding, the Cys(134) becomes more exposed but the residue is no longer capable of forming a stable disulfide in the dimeric IIC domain. In combination with the recently obtained projection structure of the IIC domain in two-dimensional crystals, a first proposal is made for alpha-helix packing in the mannitol permease. |
| Databáze: | OpenAIRE |
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