Structure of Chlorobium tepidum sepiapterin reductase complex reveals the novel substrate binding mode for stereospecific production of L-threo-tetrahydrobiopterin.
Sepiapterin reductase (SR) is involved in the last step of tetrahydrobiopterin (BH(4)) biosynthesis by reducing the di-keto group of 6-pyruvoyl tetrahydropterin. Chlorobium tepidum SR (cSR) generates a distinct BH(4) product, L-threo-BH(4) (6R-(1'S,2'S)-5,6,7,8-BH(4)), whereas animal enzymes produce L-erythro-BH(4) (6R-(1'R,2'S)-5,6,7,8-BH(4)) although it has high amino acid sequence similarities to the other animal enzymes. To elucidate the structural basis for the different reaction stereospecificities, we have determined the three-dimensional structures of cSR alone and complexed with NADP and sepiapterin at 2.1 and 1.7 A resolution, respectively. The overall folding of the cSR, the binding site for the cofactor NADP(H), and the positions of active site residues were quite similar to the mouse and the human SR. However, significant differences were found in the substrate binding region of the cSR. In comparison to the mouse SR complex, the sepiapterin in the cSR is rotated about 180 degrees around the active site and bound between two aromatic side chains of Trp-196 and Phe-99 so that its pterin ring is shifted to the opposite side, but its side chain position is not changed. The swiveled sepiapterin binding results in the conversion of the side chain configuration, exposing the opposite face for hydride transfer from NADPH. The different sepiapterin binding mode within the conserved catalytic architecture presents a novel strategy of switching the reaction stereospecificities in the same protein fold.
Supangat
Supangat
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Division of Applied Life Science, Environmental Biotechnology National Core Research Center, Gyeongsang National University, Jinju 660-701, Korea.
Seo
Kyung Hye
KH
Choi
Yong Kee
YK
Park
Young Shik
YS
Son
Daeyoung
D
Han
Chang-deok
CD
Lee
Kon Ho
KH
D000429:Alcohol Oxidoreductases / Q000737:chemistry*; D000818:Animals; D001665:Binding Sites; D001708:Biopterins / Q000031:analogs & derivatives* / Q000737:chemistry; D020134:Catalytic Domain; D041883:Chlorobium / Q000201:enzymology*; D003001:Cloning, Molecular; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000378:metabolism; D051379:Mice; D008956:Models, Chemical; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009249:NADP / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D013237:Stereoisomerism
D001708:Biopterins; D009249:NADP; D000429:Alcohol Oxidoreductases; C022630:sepiapterin reductase; C003402:sapropterin