1XWF

K185N mutated S-adenosylhomocysteine hydrolase

1XWF の概要

• エントリーDOI: 10.2210/pdb1xwf/pdb
• 分子名称: Adenosylhomocysteinase, NICOTINAMIDE-ADENINE-DINUCLEOTIDE, ADENOSINE, ... (4 entities in total)
• 機能のキーワード: s-adenosylhomocysteine hydrolase, adohcy hydrolase, adohcyase, sahh, hydrolase
• 由来する生物種: Rattus norvegicus (Norway rat)
• 細胞内の位置: Cytoplasm: P10760
• タンパク質・核酸の鎖数: 4
• 化学式量合計: 193525.20

構造登録者

Yamada, T.,Takata, Y.,Komoto, J.,Gomi, T.,Ogawa, H.,Fujioka, M.,Takusagawa, F. (登録日: 2004-11-01, 公開日: 2005-09-20, 最終更新日: 2024-02-14)

主引用文献

Yamada, T.,Takata, Y.,Komoto, J.,Gomi, T.,Ogawa, H.,Fujioka, M.,Takusagawa, F.
Catalytic mechanism of S-adenosylhomocysteine hydrolase: Roles of His 54, Asp130, Glu155, Lys185, and Aspl89.
Int.J.Biochem.Cell Biol., 37:2417-2435, 2005

PubMed Abstract: S-adenosylhomocysteine hydrolase (AdoHcyase) catalyzes the hydrolysis of S-adenosylhomocysteine (AdoHcy) to form adenosine and homocysteine. The crystal structure of the K185N mutated enzyme, which has weak catalytic activity (0.1%), has been determined at 2.8 A resolution and supports the previously predicted mechanism [Takata, Y., Yamada, T., Huang, Y., Komoto, J., Gomi, T., Ogawa, H., Fujioka, M., & Takusagawa, F. (2002). Catalytic mechanism of S-adenosylhomocysteine hydrolase. Site-directed mutagenesis of Asp-130, Lys-185, Asp-189, and Asn-190. J. Biol. Chem. 277, 22670-22676]. The mutated enzyme has an intermediate structure between the open and closed conformation, observed in the substrate-free enzyme and in the inhibitor complexes, respectively. H54, H300, and H352 were mutated to asparagine, respectively, to identify the roles of the histidine residues in catalysis. The kinetic data of H54N, H300N, and H354N mutated enzymes suggest that H54 is the amino acid residue that acts as a general acid/base to cleave the C5'-S(D) bond of AdoHcy. The E155Q mutated enzyme retained a large portion of the catalytic activity (31%), while the E155D mutated enzyme lost most of it (0.3%). The NADH accumulation measurements of the mutated enzymes indicated that the C3'-oxidation and the C4'-proton abstraction are a concerted event and the C5'-S(D) bond cleavage is an independent event. The C4'-proton exchange measurements indicate that the enzyme has an open conformation when AdoHcy is converted to 3'-keto-4', 5'-dehydro-Ado in the active site. With the results of this study and those of the previous studies, a detailed catalytic mechanism of AdoHcyase is described. K185 facilitates the C3'-oxidation, D130 abstracts the C4'-proton, D189, and E155 act as a communicator between the concerted C3'-oxidation and C4'-proton abstraction, and H54 plays as a general acid to cleave the C5'-S(D) bond of AdoHcy.

PubMed: 16061414
DOI: 10.1016/j.biocel.2005.06.009

実験手法

X-RAY DIFFRACTION (2.8 Å)