• Theresa A. Ramelot, Miami University, US
• Matthew J. Smola, Miami University, US
• Hsiau-Wei Lee, University of Georgia, US
• Colleen Ciccosanti, Rutgers University, US
• Keith Hamilton, Rutgers University, US
• Thomas B. Acton, Rutgers University, US
• Rong Xiao, Rurgers University, US
• John K. Everett, Rutgers University, US
• James H. Prestegard, University of Georgia, US
• Gaetano T. Montelione, Rutgers University, US
• Michael A. Kennedy, Miami University, US

Abstract from publication:

GmACP3 from Geobacter metallireducens is a specialized acyl carrier protein (ACP) whose gene, gmet_2339, is located near genes encoding many proteins involved in lipopolysaccharide (LPS) biosynthesis, indicating a likely function for GmACP3 in LPS production. By overexpression in Escherichia coli, about 50% holo-GmACP3 and 50% apo-GmACP3 were obtained. Apo-GmACP3 exhibited slow precipitation and non-monomeric behavior by ¹⁵N NMR relaxation measurements. Addition of 4′-phosphopantetheine (4′-PP) via enzymatic conversion by E. coli holo-ACP synthase resulted in stable >95% holo-GmACP3 that was characterized as monomeric by ¹⁵N relaxation measurements and had no indication of conformational exchange. We have determined a high-resolution solution structure of holo-GmACP3 by standard NMR methods, including refinement with two sets of NH residual dipolar couplings, allowing for a detailed structural analysis of the interactions between 4′-PP and GmACP3. Whereas the overall four helix bundle topology is similar to previously solved ACP structures, this structure has unique characteristics, including an ordered 4′-PP conformation that places the thiol at the entrance to a central hydrophobic cavity near a conserved hydrogen-bonded Trp-His pair. These residues are part of a conserved WDSLxH/N motif found in GmACP3 and its orthologs. The helix locations and the large hydrophobic cavity are more similar to medium- and long-chain acyl-ACPs than to other apo- and holo-ACP structures. Taken together, structural characterization along with bioinformatic analysis of nearby genes suggests that GmACP3 is involved in lipid A acylation, possibly by atypical long-chain hydroxy fatty acids, and potentially is involved in synthesis of secondary metabolites.

• Polypeptide chain 1
  86 residues
  PTLDALTPIFRQVFDDDSIVLTRETSANDIDAWDSLSHMNLIVSLEVHYK
  IKFALGELQKLKNVGDLADLVDKKLARKLEHHHHHH