TY - JOUR
T1 - Crystal Structure of d-Psicose 3-epimerase from Agrobacterium tumefaciens and its Complex with True Substrate d-Fructose
T2 - A Pivotal Role of Metal in Catalysis, an Active Site for the Non-phosphorylated Substrate, and its Conformational Changes
AU - Kim, Kwangsoo
AU - Kim, Hye Jung
AU - Oh, Deok Kun
AU - Cha, Sun Shin
AU - Rhee, Sangkee
N1 - Funding Information:
This work was supported by the Brain Korea 21 project, Korea Research Foundation Grant (KRF-2004-005-F00055), and by a grant (CG2114) from Crop Functional Genomics Center of the 21st Century Frontier Research Program funded by the Ministry of Science and Technology, Republic of Korea.
PY - 2006/9/1
Y1 - 2006/9/1
N2 - d-Psicose, a rare sugar produced by the enzymatic reaction of d-tagatose 3-epimerase (DTEase), has been used extensively for the bioproduction of various rare carbohydrates. Recently characterized d-psicose 3-epimerase (DPEase) from Agrobacterium tumefaciens was found to belong to the DTEase family and to catalyze the interconversion of d-fructose and d-psicose by epimerizing the C-3 position, with marked efficiency for d-psicose. The crystal structures of DPEase and its complex with the true substrate d-fructose were determined; DPEase is a tetramer and each monomer belongs to a TIM-barrel fold. The active site in each subunit is distinct from that of other TIM-barrel enzymes, which use phosphorylated ligands as the substrate. It contains a metal ion with octahedral coordination to two water molecules and four residues that are absolutely conserved across the DTEase family. Upon binding of d-fructose, the substrate displaces water molecules in the active site, with a conformation mimicking the intermediate cis-enediolate. Subsequently, Trp112 and Pro113 in the β4-α4 loop undergo significant structural changes, sealing off the active site. Structural evidence and site-directed mutagenesis of the putative catalytic residues suggest that the metal ion plays a pivotal role in catalysis by anchoring the bound d-fructose, and Glu150 and Glu244 carry out an epimerization reaction at the C-3 position.
AB - d-Psicose, a rare sugar produced by the enzymatic reaction of d-tagatose 3-epimerase (DTEase), has been used extensively for the bioproduction of various rare carbohydrates. Recently characterized d-psicose 3-epimerase (DPEase) from Agrobacterium tumefaciens was found to belong to the DTEase family and to catalyze the interconversion of d-fructose and d-psicose by epimerizing the C-3 position, with marked efficiency for d-psicose. The crystal structures of DPEase and its complex with the true substrate d-fructose were determined; DPEase is a tetramer and each monomer belongs to a TIM-barrel fold. The active site in each subunit is distinct from that of other TIM-barrel enzymes, which use phosphorylated ligands as the substrate. It contains a metal ion with octahedral coordination to two water molecules and four residues that are absolutely conserved across the DTEase family. Upon binding of d-fructose, the substrate displaces water molecules in the active site, with a conformation mimicking the intermediate cis-enediolate. Subsequently, Trp112 and Pro113 in the β4-α4 loop undergo significant structural changes, sealing off the active site. Structural evidence and site-directed mutagenesis of the putative catalytic residues suggest that the metal ion plays a pivotal role in catalysis by anchoring the bound d-fructose, and Glu150 and Glu244 carry out an epimerization reaction at the C-3 position.
KW - Agrobacterium tumefaciens
KW - crystal structure
KW - d-psicose 3-epimerase
KW - metal-dependent catalysis
KW - mutagenesis
UR - http://www.scopus.com/inward/record.url?scp=33746929133&partnerID=8YFLogxK
U2 - 10.1016/j.jmb.2006.06.069
DO - 10.1016/j.jmb.2006.06.069
M3 - Article
C2 - 16876192
AN - SCOPUS:33746929133
SN - 0022-2836
VL - 361
SP - 920
EP - 931
JO - Journal of Molecular Biology
JF - Journal of Molecular Biology
IS - 5
ER -