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The three-dimensional crystal structure of the PrpF protein of Shewanella oneidensis complexed with trans-aconitate: insights into its biological function.

Abstract
In bacteria, the dehydration of 2-methylcitrate to yield 2-methylaconitate in the 2-methylcitric acid cycle is catalyzed by a cofactor-less (PrpD) enzyme or by an aconitase-like (AcnD) enzyme. Bacteria that use AcnD also require the function of the PrpF protein, whose function was previously unknown. To gain insights into the function of PrpF, the three-dimensional crystal structure of the PrpF protein from the bacterium Shewanella oneidensis was solved at 2.0 A resolution. The protein fold of PrpF is strikingly similar to those of the non-PLP-dependent diaminopimelate epimerase from Haemophilus influenzae, a putative proline racemase from Brucella melitensis, and to a recently deposited structure of a hypothetical protein from Pseudomonas aeruginosa. Results from in vitro studies show that PrpF isomerizes trans-aconitate to cis-aconitate. It is proposed that PrpF catalysis of the cis-trans isomerization proceeds through a base-catalyzed proton abstraction coupled with a rotation about C2-C3 bond of 2-methylaconitate, and that residue Lys73 is critical for PrpF function. The newly identified function of PrpF as a non-PLP-dependent isomerase, together with the fact that PrpD-containing bacteria do not require PrpF, suggest that the isomer of 2-methylaconitate that serves as a substrate of aconitase must have the same stereochemistry as that synthesized by PrpD. From this, it follows that the 2-methylaconitate isomer generated by AcnD is not a substrate of aconitase, and that PrpF is required to generate the correct isomer. As a consequence, the isomerase activity of PrpF may now be viewed as an integral part of the 2-methylcitric acid cycle.
AuthorsGraeme S Garvey, Christopher J Rocco, Jorge C Escalante-Semerena, Ivan Rayment
JournalProtein science : a publication of the Protein Society (Protein Sci) Vol. 16 Issue 7 Pg. 1274-84 (Jul 2007) ISSN: 0961-8368 [Print] United States
PMID17567742 (Publication Type: Journal Article, Research Support, N.I.H., Extramural)
Chemical References
  • Bacterial Proteins
  • Citrates
  • 2-methylcitric acid
  • Aconitic Acid
  • Hydro-Lyases
  • Aconitate Hydratase
Topics
  • Aconitate Hydratase (chemistry, metabolism)
  • Aconitic Acid (chemistry, metabolism)
  • Bacterial Proteins (chemistry, metabolism)
  • Citrates (chemistry, metabolism)
  • Crystallography, X-Ray
  • Dimerization
  • Hydro-Lyases (chemistry, metabolism)
  • Isomerism
  • Models, Biological
  • Models, Molecular
  • Molecular Structure
  • Protein Binding
  • Protein Conformation
  • Protein Structure, Tertiary
  • Shewanella (metabolism)

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