The structural basis of Erwinia rhapontici isomaltulose synthase.
Sucrose isomerase NX-5 from Erwiniarhapontici efficiently catalyzes the isomerization of sucrose to isomaltulose (main product) and trehalulose (by-product). To investigate the molecular mechanism controlling sucrose isomer formation, we determined the crystal structures of native NX-5 and its mutant complexes E295Q/sucrose and D241A/glucose at 1.70 Å, 1.70 Å and 2.00 Å, respectively. The overall structure and active site architecture of NX-5 resemble those of other reported sucrose isomerases. Strikingly, the substrate binding mode of NX-5 is also similar to that of trehalulose synthase from Pseudomonasmesoacidophila MX-45 (MutB). Detailed structural analysis revealed the catalytic RXDRX motif and the adjacent 10-residue loop of NX-5 and isomaltulose synthase PalI from Klebsiella sp. LX3 adopt a distinct orientation from those of trehalulose synthases. Mutations of the loop region of NX-5 resulted in significant changes of the product ratio between isomaltulose and trehalulose. The molecular dynamics simulation data supported the product specificity of NX-5 towards isomaltulose and the role of the loop(330-339) in NX-5 catalysis. This work should prove useful for the engineering of sucrose isomerase for industrial carbohydrate biotransformations.
State Key Laboratory of Materials-Oriented Chemical Engineering, College of Food Science and Light Industry, Nanjing University of Technology, Nanjing, China ; State Key Laboratory of Bio-Organic and Natural Products Chemistry, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, Shanghai, China.
Zheng
Z
Xu
Sha
S
Li
Jie
J
Li
Yan
Y
Li
Xiaohai
X
Feng
Renxiao
R
Wang
Hong
H
Xu
Jiahai
J
Zhou
D000595:Amino Acid Sequence; D020134:Catalytic Domain; D004885:Erwinia / Q000201:enzymology* / Q000235:genetics; D005947:Glucose / Q000737:chemistry / Q000378:metabolism; D019751:Intramolecular Transferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D007536:Isomerism; D008958:Models, Molecular; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D011485:Protein Binding; D011487:Protein Conformation; D016415:Sequence Alignment; D013395:Sucrose / Q000737:chemistry / Q000378:metabolism
D013395:Sucrose; D019751:Intramolecular Transferases; C042002:isomaltulose synthase; D005947:Glucose