title: Differential repositioning of the second transmembrane helices from E. coli Tar and EnvZ upon moving the flanking aromatic residues.
abstract: Aromatic tuning, i.e. repositioning aromatic residues found at the cytoplasmic end of transmembrane (TM) domains within bacterial receptors, has been previously shown to modulate signal output from the aspartate chemoreceptor (Tar) and the major osmosensor EnvZ of Escherichia coli. In the case of Tar, changes in signal output consistent with the vertical position of the native Trp-Tyr aromatic tandem within TM2 were observed. In contrast, within EnvZ, where a Trp-Leu-Phe aromatic triplet was repositioned, the surface that the triplet resided upon was the major determinant governing signal output. However, these studies failed to determine whether moving the aromatic residues was sufficient to physically reposition the TM helix within a membrane. Recent coarse-grained molecular dynamics (CG-MD) simulations predicted displacement of Tar TM2 upon moving the aromatic residues at the cytoplasmic end of the helix. Here, we demonstrate that repositioning the Trp-Tyr tandem within Tar TM2 displaces the C-terminal boundary of the helix relative to the membrane. In a similar analysis of EnvZ, an abrupt initial displacement of TM2 was observed but no subsequent movement was seen, suggesting that the vertical position of TM2 is not governed by the location of the Trp-Leu-Phe triplet. Our results also provide another set of experimental data, i.e. the resistance of EnvZ TM2 to being displaced upon aromatic tuning, which could be useful for subsequent refinement of the initial CG-MD simulations. Finally, we discuss the limitations of these methodologies, how moving flanking aromatic residues might impact steady-state signal output and the potential to employ aromatic tuning in other bacterial membrane-spanning receptors.
authors.affiliation: Department of Biochemistry and Biophysics, Stockholm University, Svante Arrhenius väg 16C, SE-10691 Stockholm, Sweden.
authors.forename: Salomé C
authors.initials: SC
authors.lastname: Botelho
authors.affiliation: Department of Biochemistry and Biophysics, Stockholm University, Svante Arrhenius väg 16C, SE-10691 Stockholm, Sweden.
authors.forename: Karl
authors.initials: K
authors.lastname: Enquist
authors.affiliation: Department of Biochemistry and Biophysics, Stockholm University, Svante Arrhenius väg 16C, SE-10691 Stockholm, Sweden.
authors.forename: Gunnar
authors.initials: G
authors.lastname: von Heijne
authors.affiliation: Division of Pharmacy, Durham University, Queen's Campus, Stockton-on-Tees TS17 6BH, England, UK; Wolfson Research Institute for Health and Wellbeing, Durham University, Queen's Campus, Stockton-on-Tees, TS17 6BH, England, UK. Electronic address: roger.draheim@durham.ac.uk.
authors.forename: Roger R
authors.initials: RR
authors.lastname: Draheim
mesh_terms: D020816:Amino Acid Motifs; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry*; D002462:Cell Membrane / Q000737:chemistry* / Q000378:metabolism; D003593:Cytoplasm / Q000737:chemistry* / Q000378:metabolism; D004926:Escherichia coli / Q000737:chemistry* / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry*; D007930:Leucine / Q000737:chemistry; D056004:Molecular Dynamics Simulation; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000737:chemistry*; D010649:Phenylalanine / Q000737:chemistry; D017433:Protein Structure, Secondary; D011956:Receptors, Cell Surface / Q000737:chemistry*; D016415:Sequence Alignment; D015398:Signal Transduction; D013329:Structure-Activity Relationship; D014364:Tryptophan / Q000737:chemistry
chemical_list: D001425:Bacterial Outer Membrane Proteins; D029968:Escherichia coli Proteins; D009097:Multienzyme Complexes; D011956:Receptors, Cell Surface; C057331:Tar protein, E coli; D010649:Phenylalanine; D014364:Tryptophan; C057843:envZ protein, E coli; D007930:Leucine
keywords: Aromatic tuning; Glycosylation mapping; Hydrophobic-polar membrane interface; Interfacial anchoring; Transmembrane helices