Structural and catalytic properties of the D-3-hydroxybutyrate dehydrogenase from Pseudomonas aeruginosa.

Driss Mountassif, Pierre Andreoletti, Mustapha Cherkaoui-Malki, Norbert Latruffe, M'hammed Saïd El Kebbaj

Index: Curr. Microbiol. 61 , 7-12, (2010)

Full Text: HTML

Abstract

To put forward BDH from Pseudomonas aeruginosa's enzymatic properties, we report a two-step purification of BDH and its gene sequencing allowing the investigation of its structural properties. Purification of BDH was achieved, using ammonium sulfate fractionation and Blue Sepharose CL-6B affinity chromatography. SDS-PAGE analysis reveals a MM of 29 kDa, whereas the native enzyme showed a MM of 120 kDa suggesting a homotetrameric structure. BDH encoding gene sequence shows a nucleotide open reading frame sequence of 771 bp encoding a 265 amino acid residues polypeptide chain. The modeling analysis of the three dimensional structure fits with the importance of amino acids in the catalysis reaction especially a strictly conserved tetrad. Amino-acid residues in interaction with the coenzyme NAD(+) were also identified.