Unknown

Dataset Information

0

Structure and catalytic mechanism of 3-ketosteroid-Delta4-(5α)-dehydrogenase from Rhodococcus jostii RHA1 genome.


ABSTRACT: 3-Ketosteroid Δ4-(5α)-dehydrogenases (Δ4-(5α)-KSTDs) are enzymes that introduce a double bond between the C4 and C5 atoms of 3-keto-(5α)-steroids. Here we show that the ro05698 gene from Rhodococcus jostii RHA1 codes for a flavoprotein with Δ4-(5α)-KSTD activity. The 1.6 Å resolution crystal structure of the enzyme revealed three conserved residues (Tyr-319, Tyr-466, and Ser-468) in a pocket near the isoalloxazine ring system of the FAD co-factor. Site-directed mutagenesis of these residues confirmed that they are absolutely essential for catalytic activity. A crystal structure with bound product 4-androstene-3,17-dione showed that Ser-468 is in a position in which it can serve as the base abstracting the 4β-proton from the C4 atom of the substrate. Ser-468 is assisted by Tyr-319, which possibly is involved in shuttling the proton to the solvent. Tyr-466 is at hydrogen bonding distance to the C3 oxygen atom of the substrate and can stabilize the keto-enol intermediate occurring during the reaction. Finally, the FAD N5 atom is in a position to be able to abstract the 5α-hydrogen of the substrate as a hydride ion. These features fully explain the reaction catalyzed by Δ4-(5α)-KSTDs.

SUBMITTER: van Oosterwijk N 

PROVIDER: S-EPMC3438930 | BioStudies | 2012-01-01

SECONDARY ACCESSION(S): 1d4c

REPOSITORIES: biostudies

Similar Datasets

2013-01-01 | S-EPMC3853301 | BioStudies
2001-01-01 | S-EPMC1222170 | BioStudies
1995-01-01 | S-EPMC1136746 | BioStudies
2007-01-01 | S-EPMC1851095 | BioStudies
2015-01-01 | S-EPMC4310174 | BioStudies
2013-01-01 | S-EPMC3648074 | BioStudies
2008-01-01 | S-EPMC2276324 | BioStudies
2009-01-01 | S-EPMC2794732 | BioStudies
2007-01-01 | S-EPMC2222818 | BioStudies
2008-01-01 | S-EPMC2569941 | BioStudies