title: Solution structure of the C-terminal antiparallel coiled-coil domain from Escherichia coli osmosensor ProP.
abstract: Bacteria respond to increasing medium osmolality by accumulating organic solutes that are compatible with cellular functions. Transporter ProP of Escherichia coli, a proton symporter and a member of the major facilitator superfamily, senses osmotic shifts and responds by importing osmolytes such as glycine betaine. ProP contains a cytoplasmic, C-terminal extension that is essential for its activity. A peptide corresponding to the C-terminal extension of ProP forms a homodimeric alpha-helical coiled-coil even though some of its heptad a positions are not occupied by hydrophobic amino acid residues. Unexpectedly, amino acid replacement R488I, occurring at a heptad a position, destabilized the coiled-coil formed by the ProP peptide and attenuated the response of the intact transporter to osmotic upshifts in vivo. Thus, ProP was proposed to dimerize via an antiparallel coiled-coil. We used nuclear magnetic resonance (NMR) spectroscopy to determine the structure of the synthetic peptide corresponding to residues 468-497 of ProP. This region did form an antiparallel coil-coil in which critical residue R488 specifies the antiparallel coiled-coil orientation by forming stabilizing salt-bridges. Charged residues (both acidic and basic) are clustered on the c/g surface of the coiled-coil whereas polar residues are distributed on the b/e surface. This causes the structure to be bent, in contrast to other known antiparallel coiled-coils (those from the hepatitis delta antigen (PDB ID code 1A92) and the bovine F(1) ATPase inhibitor protein (PDB ID code 1HF9)). The coiled-coil and its possible importance for osmosensing are discussed.
authors.affiliation: Department of Biochemistry and Molecular Genetics, University of Colorado Health Sciences Center, Denver, CO 80262, USA.
authors.forename: David L
authors.initials: DL
authors.lastname: Zoetewey
authors.forename: Brian P
authors.initials: BP
authors.lastname: Tripet
authors.forename: Tatiana G
authors.initials: TG
authors.lastname: Kutateladze
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Overduin
authors.forename: Janet M
authors.initials: JM
authors.lastname: Wood
authors.forename: Robert S
authors.initials: RS
authors.lastname: Hodges
mesh_terms: D000595:Amino Acid Sequence; D004926:Escherichia coli / Q000737:chemistry*; D029968:Escherichia coli Proteins / Q000737:chemistry*; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D011487:Protein Conformation; D017386:Sequence Homology, Amino Acid; D012996:Solutions; D027981:Symporters / Q000737:chemistry*
chemical_list: D029968:Escherichia coli Proteins; C079244:ProP protein, E coli; D012996:Solutions; D027981:Symporters