Crystallization of 4-hydroxybenzoyl-CoA reductase and the structure of its electron donor ferredoxin

Acta Crystallogr D Biol Crystallogr. 2004 Feb;60(Pt 2):388-91. doi: 10.1107/S0907444903028506. Epub 2004 Jan 23.

Abstract

4-Hydroxybenzoyl-CoA reductase (4-HBCR) is a central enzyme in the metabolism of phenolic compounds in anaerobic bacteria. The enzyme catalyzes the reductive removal of the phenolic hydroxyl group from 4-hydroxybenzoyl-CoA, yielding benzoyl-CoA and water. 4-HBCR belongs to the xanthine oxidase (XO) family of molybdenum enzymes which occur as heterodimers, (alphabetagamma)(2). 4-HBCR contains two molybdopterins, four [2Fe-2S] and two [4Fe-4S] clusters and two FADs. A low-potential Allochromatium vinosum-type ferredoxin containing two [4Fe-4S] clusters serves as an in vivo electron donor for 4-HBCR. In this work, the oxygen-sensitive proteins 4-HBCR and the ferredoxin (TaFd) from Thauera aromatica were crystallized under anaerobic conditions. 4-HBCR crystallized with PEG 4000 and MPD as precipitant diffracted to about 1.6 A resolution and the crystals were highly suitable for X-ray structure analysis. Crystals of TaFd were obtained with (NH(4))(3)PO(4) as precipitant and revealed a solvent content of 77%, which is remarkably high for a small soluble protein. The structure of TaFd was solved at 2.9 A resolution by the molecular-replacement method using the highly related structure of the ferredoxin (CvFd) from A. vinosum as a model. Structural changes between the two ferredoxins around the [4Fe-4S] cluster can be correlated with their different redox potentials.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Crystallography, X-Ray / methods*
  • Ferredoxins / chemistry
  • Models, Chemical
  • Models, Molecular
  • Oxidation-Reduction
  • Oxidoreductases Acting on CH-CH Group Donors / chemistry*
  • Protein Conformation
  • Protein Structure, Tertiary
  • Thauera / enzymology*
  • Xanthine Oxidase / chemistry

Substances

  • Ferredoxins
  • Xanthine Oxidase
  • Oxidoreductases Acting on CH-CH Group Donors
  • benzoyl-coenzyme A-4-oxidoreductase