Crystallization and preliminary X-ray analysis of dmpFG-encoded 4-hydroxy-2-ketovalerate aldolase--aldehyde dehydrogenase (acylating) from Pseudomonas sp. strain CF600.
The final two steps of the meta-cleavage pathway for catechol degradation in Pseudomonas sp. strain CF600 involve the conversion of 4-hydroxy-2-ketovalerate to pyruvate and acetyl coenzyme A by the enzymes 4-hydroxy-2-ketovalerate aldolase and NAD(+)-dependent acylating aldehyde dehydrogenase. Biochemical studies indicate that these two enzymes comprise a bifunctional heterodimer (DmpFG, molecular mass 71 kDa) and suggest that the product of the aldolase reaction is transferred to the dehydrogenase active site via a channeling mechanism. Crystals of the DmpFG complex grow in multiple fan-like clusters of thin plates by the hanging-drop method and are improved by streak-seeding. The crystals belong to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 102.0, b = 140.7, c = 191.3 A, and diffract to 2.1 A resolution. The asymmetric unit contains four DmpFG heterodimers. Heavy-atom derivative screening identified three isomorphous derivatives.
Manjasetty
B A
BA
Department of Physics, Government Science College, Bangalore 560 012, India.
Croteau
N
N
Powlowski
J
J
Vrielink
A
A
D000445:Aldehyde Oxidoreductases / Q000737:chemistry* / Q000378:metabolism; D001665:Binding Sites; D003460:Crystallization; D019281:Dimerization; D009097:Multienzyme Complexes / Q000737:chemistry* / Q000378:metabolism; D009243:NAD / Q000378:metabolism; D007652:Oxo-Acid-Lyases / Q000737:chemistry* / Q000378:metabolism; D011549:Pseudomonas / Q000201:enzymology*; D017386:Sequence Homology, Amino Acid; D012984:Software; D014961:X-Ray Diffraction / Q000379:methods
D009097:Multienzyme Complexes; D009243:NAD; D000445:Aldehyde Oxidoreductases; C027014:acetaldehyde dehydrogenase (acylating); C038482:4-hydroxy-2-ketovalerate aldolase; D007652:Oxo-Acid-Lyases