Structural basis of the sphingomyelin phosphodiesterase activity in neutral sphingomyelinase from Bacillus cereus.
Sphingomyelinase (SMase) from Bacillus cereus (Bc-SMase) hydrolyzes sphingomyelin to phosphocholine and ceramide in a divalent metal ion-dependent manner. Bc-SMase is a homologue of mammalian neutral SMase (nSMase) and mimics the actions of the endogenous mammalian nSMase in causing differentiation, development, aging, and apoptosis. Thus Bc-SMase may be a good model for the poorly characterized mammalian nSMase. The metal ion activation of sphingomyelinase activity of Bc-SMase was in the order Co2+ > or = Mn2+ > or = Mg2+ >> Ca2+ > or = Sr2+. The first crystal structures of Bc-SMase bound to Co2+, Mg2+, or Ca2+ were determined. The water-bridged double divalent metal ions at the center of the cleft in both the Co2+- and Mg2+-bound forms were concluded to be the catalytic architecture required for sphingomyelinase activity. In contrast, the architecture of Ca2+ binding at the site showed only one binding site. A further single metal-binding site exists at one side edge of the cleft. Based on the highly conserved nature of the residues of the binding sites, the crystal structure of Bc-SMase with bound Mg2+ or Co2+ may provide a common structural framework applicable to phosphohydrolases belonging to the DNase I-like folding superfamily. In addition, the structural features and site-directed mutagenesis suggest that the specific beta-hairpin with the aromatic amino acid residues participates in binding to the membrane-bound sphingomyelin substrate.
Ago
Hideo
H
Structural Biophysics Laboratory, RIKEN SPring-8 Center, Harima Institute, 1-1-1 Kouto, Sayo, Hyogo 679-5148, Japan. ago@spring8.or.jp
Oda
Masataka
M
Takahashi
Masaya
M
Tsuge
Hideaki
H
Ochi
Sadayuki
S
Katunuma
Nobuhiko
N
Miyano
Masashi
M
Sakurai
Jun
J
D000595:Amino Acid Sequence; D001409:Bacillus cereus / Q000201:enzymology*; D001483:Base Sequence; D001665:Binding Sites; D017931:DNA Primers; D008670:Metals / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D017386:Sequence Homology, Amino Acid; D013108:Sphingomyelin Phosphodiesterase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014961:X-Ray Diffraction
D017931:DNA Primers; D008670:Metals; D013108:Sphingomyelin Phosphodiesterase