Structural and mechanistic insights into the kynurenine aminotransferase-mediated excretion of kynurenic acid.
Kynurenine aminotransferase (KAT) is a homodimeric pyridoxal protein that mediates the catalytic conversion of kynurenine (KYN) to kynurenic acid (KYA), an endogenous N-methyl-d-aspartate (NMDA) receptor antagonist. KAT is involved in the biosynthesis of glutamic and aspartic acid, functions as a neurotransmitter for the NMDA receptor in mammals, and is regulated by allosteric mechanisms. Its importance in various diseases such as schizophrenia makes KAT a highly attractive drug target. Here, we present the crystal structure of the Pyrococcus horikoshii KAT (PhKAT) in complex with pyridoxamine phosphates (PMP), KYN, and KYA. Surprisingly, the PMP was bound to the LYS-269 of phKAT by forming a covalent hydrazine bond. This crystal structure clearly shows that an amino group of KYN was transaminated to PLP, which forms a Schiff's base with the LYS-269 of the KYN. Thus, our structure confirms that the PMPs represent an intermediate state during the KAT reaction. Thus, PhKAT catalyzes the sequential conversion of KYN to KYA via the formation of an intermediate 4-(2-aminophenyl)-2,4-dioxobutanoate (4AD), which is spontaneously converted to KYA in the absence of an amino group acceptor. Furthermore, we identified the two entry and exit sites of the PhKAT homodimer for KYN and KYA, respectively. The structural data on PhKAT presented in this manuscript contributes to further the understanding of transaminase enzyme reaction mechanisms.
Department of Material and Life Science, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan; Department of Drug Discovery, Center for Development of Advanced Medicine for Dementia, National Center for Geriatrics and Gerontology, 35 Gengo, Morioka, Obu, Aichi 474-8511, Japan. Electronic address: okada@ncgg.go.jp.
Ken
K
Okada
Department of Material and Life Science, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan; International College, Osaka University, 1-30 Machikaneyama-cho, Toyonaka, Osaka 560-0043, Japan.
Clement
C
Angkawidjaja
Department of Material and Life Science, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan.
Yuichi
Y
Koga
Department of Material and Life Science, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan.
Shigenori
S
Kanaya
D000595:Amino Acid Sequence; D019843:Archaeal Proteins / Q000737:chemistry / Q000378:metabolism; D007736:Kynurenic Acid / Q000737:chemistry / Q000378:metabolism*; D007737:Kynurenine / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D011733:Pyridoxamine / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D043902:Pyrococcus horikoshii / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D000637:Transaminases / Q000737:chemistry* / Q000378:metabolism*
D019843:Archaeal Proteins; D007737:Kynurenine; D011733:Pyridoxamine; D000637:Transaminases; C021481:kynurenine-oxoglutarate transaminase; D007736:Kynurenic Acid; C010627:pyridoxamine phosphate
Excretion mechanism; Kynurenic acid; Kynurenine aminotransferase; Pyridoxamine phosphate