Solution structure of the C-terminal antiparallel coiled-coil domain from Escherichia coli osmosensor ProP

• Published in: Journal of molecular biology Vol. 334; no. 5; pp. 1063 - 1076
• Main Authors: Zoetewey, David L, Tripet, Brian P, Kutateladze, Tatiana G, Overduin, Michael J, Wood, Janet M, Hodges, Robert S
• Format: Journal Article
• Language: English
• Published: England 12.12.2003
• Subjects: Amino Acid Sequence; Escherichia coli; Escherichia coli - chemistry; Escherichia coli Proteins - chemistry; Molecular Sequence Data; Nuclear Magnetic Resonance, Biomolecular; Protein Conformation; Sequence Homology, Amino Acid; Solutions; Symporters - chemistry
• ISSN: 0022-2836; 1089-8638
• DOI: 10.1016/j.jmb.2003.10.020

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.