Octomeric pyruvate-ferredoxin oxidoreductase from Desulfovibrio vulgaris.
Pyruvate-ferredoxin oxidoreductatse (PFOR) carries out the central step in oxidative decarboxylation of pyruvate to acetyl-CoA. We have purified this enzyme from Desulfovibrio vulgaris Hildenborough (DvH) as part of a systematic characterization of as many multiprotein complexes as possible for this organism, and the three-dimensional structure of this enzyme has been determined by a combination of electron microscopy (EM), single particle image analysis, homology modeling and computational molecular docking. Our results show that the 1MDa DvH PFOR complex is a homo-octomer, or more precisely, a tetramer of the dimeric form of the related enzyme found in Desulfovibrio africanus (Da), with which it shares a sequence identity of 69%. Our homology model of the DvH PFOR dimer is based on the Da PFOR X-ray structure. Docking of this model into our 17A resolution EM-reconstruction of negatively stained DvH PFOR octomers strongly suggests that the difference in oligomerization state for the two species is due to the insertion of a single valine residue (Val383) within a surface loop of the DvH enzyme. This study demonstrates that the strategy of intermediate resolution EM reconstruction coupled to homology modeling and docking can be powerful enough to infer the functionality of single amino acid residues.
Garczarek
Florian
F
Life Sciences Division, Lawrence Berkeley National Laboratory, University of California, Berkeley, CA 94720, USA.
Dong
Ming
M
Typke
Dieter
D
Witkowska
H Ewa
HE
Hazen
Terry C
TC
Nogales
Eva
E
Biggin
Mark D
MD
Glaeser
Robert M
RM
D000596:Amino Acids; D019295:Computational Biology; D016969:Desulfovibrio vulgaris / Q000201:enzymology*; D019281:Dimerization; D008854:Microscopy, Electron; D008958:Models, Molecular; D011487:Protein Conformation; D050743:Pyruvate Synthase / Q000737:chemistry* / Q000302:isolation & purification; D014633:Valine
D000596:Amino Acids; D050743:Pyruvate Synthase; D014633:Valine