Crystal structure of CYP105A1 (P450SU-1) in complex with 1alpha,25-dihydroxyvitamin D3.
Vitamin D 3 (VD 3), a prohormone in mammals, plays a crucial role in the maintenance of calcium and phosphorus concentrations in serum. Activation of VD 3 requires 25-hydroxylation in the liver and 1alpha-hydroxylation in the kidney by cytochrome P450 (CYP) enzymes. Bacterial CYP105A1 converts VD 3 into 1alpha,25-dihydroxyvitamin D 3 (1alpha,25(OH) 2D 3) in two independent reactions, despite its low sequence identity with mammalian enzymes (<21% identity). The present study determined the crystal structures of a highly active mutant (R84A) of CYP105A1 from Streptomyces griseolus in complex and not in complex with 1alpha,25(OH) 2D 3. The compound 1alpha,25(OH) 2D 3 is positioned 11 A from the iron atom along the I helix within the pocket. A similar binding mode is observed in the structure of the human CYP2R1-VD 3 complex, indicating a common substrate-binding mechanism for 25-hydroxylation. A comparison with the structure of wild-type CYP105A1 suggests that the loss of two hydrogen bonds in the R84A mutant increases the adaptability of the B' and F helices, creating a transient binding site. Further mutational analysis of the active site reveals that 25- and 1alpha-hydroxylations share residues that participate in these reactions. These results provide the structural basis for understanding the mechanism of the two-step hydroxylation that activates VD 3.
Sugimoto
Hiroshi
H
RIKEN SPring-8 Center, Harima Institute, Sayo, Hyogo 679-5148, Japan. sugimoto@spring8.or.jp
Shinkyo
Raku
R
Hayashi
Keiko
K
Yoneda
Sachiyo
S
Yamada
Masato
M
Kamakura
Masaki
M
Ikushiro
Shin-ichi
S
Shiro
Yoshitsugu
Y
Sakaki
Toshiyuki
T
D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D001483:Base Sequence; D001665:Binding Sites; D002117:Calcitriol / Q000737:chemistry*; D018360:Crystallography, X-Ray; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry* / Q000235:genetics; D017931:DNA Primers; D008958:Models, Molecular; D008968:Molecular Conformation; D016297:Mutagenesis, Site-Directed; D010105:Oxygenases / Q000737:chemistry* / Q000235:genetics; D016133:Polymerase Chain Reaction
D001426:Bacterial Proteins; D017931:DNA Primers; D003577:Cytochrome P-450 Enzyme System; D010105:Oxygenases; C086254:P450SU1 protein, Streptomyces griseolus; D002117:Calcitriol