Function of the N-terminal cap of the PAS domain in signaling by the aerotaxis receptor Aer.
Aer, the Escherichia coli receptor for behavioral responses to oxygen (aerotaxis), energy, and redox potential, contains a PAS sensory-input domain. Within the PAS superfamily, the N-terminal segment (N-cap) is poorly conserved and its role is not well understood. We investigated the role of the N-cap (residues 1 to 19) in the Aer PAS domain by missense and truncation mutagenesis. Aer-PAS N-cap truncations and an Aer-M21P substitution resulted in low cellular levels of the mutant proteins, suggesting that the N-terminal region was important for stabilizing the structure of the PAS domain. The junction of the N-cap and PAS core was critical for signaling in Aer. Mutations and truncations in the sequence encoding residues 15 to 21 introduced a range of phenotypes, including defects in FAD binding, constant tumbling motility, and an inverse response in which E. coli cells migrated away from oxygen concentrations to which they are normally attracted. The proximity of two N-cap regions in an Aer dimer was assessed in vivo by oxidatively cross-linking serial cysteine substitutions. Cross-linking of several cysteine replacements at 23 degrees C was attenuated at 10 degrees C, indicating contact was not at a stable dimer interface but required lateral mobility. We observed large multimers of Aer when we combined cross-linking of N-cap residues with a cysteine replacement that cross-links exclusively at the Aer dimer interface. This suggests that the PAS N-cap faces outwards in a dimer and that PAS-PAS contacts can occur between adjacent dimers.
Watts
Kylie J
KJ
Division of Microbiology and Molecular Genetics, Loma Linda University, Loma Linda, CA 92350, USA.
Sommer
Kirsten
K
Fry
Sheena L
SL
Johnson
Mark S
MS
Taylor
Barry L
BL
D019943:Amino Acid Substitution; D002352:Carrier Proteins / Q000235:genetics / Q000502:physiology*; D002633:Chemotaxis / Q000235:genetics; D018389:Codon, Nonsense; D019281:Dimerization; D004926:Escherichia coli / Q000235:genetics / Q000502:physiology*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000502:physiology*; D005182:Flavin-Adenine Dinucleotide / Q000378:metabolism; D036341:Intercellular Signaling Peptides and Proteins; D008958:Models, Molecular; D009068:Movement; D020125:Mutation, Missense; D017434:Protein Structure, Tertiary / Q000235:genetics / Q000502:physiology; D021122:Protein Subunits / Q000378:metabolism; D017384:Sequence Deletion; D015398:Signal Transduction / Q000235:genetics
C108679:Aer protein, E coli; D002352:Carrier Proteins; D018389:Codon, Nonsense; D029968:Escherichia coli Proteins; D036341:Intercellular Signaling Peptides and Proteins; D021122:Protein Subunits; D005182:Flavin-Adenine Dinucleotide