Channeling and conformational changes in the heterotetrameric sarcosine oxidase from Corynebacterium sp. U-96.
We characterized the crystal structures of heterotetrameric sarcosine oxidase (SO) from Corynebacterium sp. U-96 complexed with methylthioacetate (MTA), pyrrole 2-carboxylate (PCA) and sulphite, and of sarcosine-reduced SO. SO comprises α-, β-, γ- and δ-subunits; FAD and FMN cofactors; and a large internal cavity. MTA and PCA are sandwiched between the re-face of the FAD isoalloxazine ring and the β-subunit C-terminal residues. Reduction of flavin cofactors shifts the β-subunit Ala1 towards the α-subunit Met55, forming a surface cavity at the oxygen-channel vestibule and rendering the β-subunit C-terminal residues mobile. We identified three channels connecting the cavity and the enzyme surface. Two of them exist in the inter-subunit space between α and β-subunits, and the substrate sarcosine seems to enter the active site through either of these channels and reaches the re-side of the FAD isoalloxazine ring by traversing the mobile β-subunit C-terminal residues. The third channel goes through the α-subunit and has a folinic acid-binding site, where the iminium intermediate is converted to Gly and either formaldehyde or, 5,10-methylenetetrahydrofolate. Oxygen molecules are probably located on the surface cavity and diffuse to the FMN isoalloxazine ring; the H(2)O(2) formed exits via the oxygen channel.
Moriguchi
Tomotaka
T
Division of Bioscience, Graduate School of Fundamental Life Science, Kitasato University, Kitasato 1-15-1, Sagamihara, Kanagawa 252-0329, Japan.
Ida
Koh
K
Hikima
Takaaki
T
Ueno
Go
G
Yamamoto
Masaki
M
Suzuki
Haruo
H
D020134:Catalytic Domain; D003352:Corynebacterium / Q000201:enzymology*; D018360:Crystallography, X-Ray; D002955:Leucovorin / Q000737:chemistry / Q000378:metabolism; D008958:Models, Molecular; D015394:Molecular Structure; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D050879:Sarcosine Oxidase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013763:Tetrahydrofolates / Q000737:chemistry / Q000378:metabolism
D021122:Protein Subunits; D013763:Tetrahydrofolates; D050879:Sarcosine Oxidase; D002955:Leucovorin