title: Quantitative approaches to utilizing mutational analysis and disulfide crosslinking for modeling a transmembrane domain.
abstract: The transmembrane domain of chemoreceptor Trg from Escherichia coli contains four transmembrane segments in its native homodimer, two from each subunit. We had previously used mutational analysis and sulfhydryl cross-linking between introduced cysteines to obtain data relevant to the three-dimensional organization of this domain. In the current study we used Fourier analysis to assess these data quantitatively for periodicity along the sequences of the segments. The analyses provided a strong indication of alpha-helical periodicity in the first transmembrane segment and a substantial indication of that periodicity for the second segment. On this basis, we considered both segments as idealized alpha-helices and proceeded to model the transmembrane domain as a unit of four helices. For this modeling, we calculated helical crosslinking moments, parameters analogous to helical hydrophobic moments, as a quantitative way of condensing and utilizing a large body of crosslinking data. Crosslinking moments were used to define the relative separation and orientation of helical pairs, thus creating a quantitatively derived model for the transmembrane domain of Trg. Utilization of Fourier transforms to provide a quantitative indication of periodicity in data from analyses of transmembrane segments, in combination with helical crosslinking moments to position helical pairs should be useful in modeling other transmembrane domains.
authors.affiliation: Department of Biochemistry and Biophysics, Washington State University, Pullman 99164-4660, USA.
authors.forename: G F
authors.initials: GF
authors.lastname: Lee
authors.forename: G L
authors.initials: GL
authors.lastname: Hazelbauer
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry*; D002633:Chemotaxis; D003432:Cross-Linking Reagents; D004252:DNA Mutational Analysis; D004220:Disulfides / Q000737:chemistry; D004926:Escherichia coli / Q000737:chemistry*; D029968:Escherichia coli Proteins; D005583:Fourier Analysis; D008565:Membrane Proteins; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry*; D010507:Periodicity; D017433:Protein Structure, Secondary; D011956:Receptors, Cell Surface / Q000737:chemistry*
chemical_list: D001426:Bacterial Proteins; D003432:Cross-Linking Reagents; D004220:Disulfides; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D010446:Peptide Fragments; D011956:Receptors, Cell Surface; C041476:Trg protein, E coli