title: Reengineering substrate specificity of E. coli glutamate dehydrogenase using a position-based prediction method.
abstract: OBJECTIVE
  To re-engineer the active site of proteins for non-natural substrates using a position-based prediction method (PBPM).
  
  
  RESULTS
  The approach has been applied to re-engineer the E. coli glutamate dehydrogenase to alter its substrate from glutamate to homoserine for a de novo 1,3-propanediol biosynthetic pathway. After identification of key residues that determine the substrate specificity, residue K92 was selected as a candidate site for mutation. Among the three mutations (K92V, K92C, and K92M) suggested by PBPM, the specific activity of the best mutant (K92 V) was increased from 171 ± 35 to 1328 ± 71 μU mg-1.
  
  
  CONCLUSIONS
  The PBPM approach has a high efficiency for re-engineering the substrate specificity of natural enzymes for new substrates.
authors.affiliation: Institute of Bioprocess and Biosystems Engineering, Hamburg University of Technology, Denickestrasse 15, 21073, Hamburg, Germany.
authors.forename: Feng
authors.initials: F
authors.lastname: Geng
authors.affiliation: Institute of Bioprocess and Biosystems Engineering, Hamburg University of Technology, Denickestrasse 15, 21073, Hamburg, Germany.
authors.forename: Cheng-Wei
authors.initials: CW
authors.lastname: Ma
authors.affiliation: Institute of Bioprocess and Biosystems Engineering, Hamburg University of Technology, Denickestrasse 15, 21073, Hamburg, Germany. aze@tuhh.de.
authors.forename: An-Ping
authors.initials: AP
authors.lastname: Zeng
mesh_terms: D020134:Catalytic Domain; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D005969:Glutamate Dehydrogenase / Q000235:genetics / Q000378:metabolism*; D018698:Glutamic Acid / Q000378:metabolism; D006714:Homoserine / Q000378:metabolism; D008958:Models, Molecular; D009154:Mutation; D011409:Propylene Glycols / Q000378:metabolism; D015202:Protein Engineering; D017434:Protein Structure, Tertiary; D013379:Substrate Specificity
chemical_list: D011409:Propylene Glycols; D018698:Glutamic Acid; C041787:1,3-propanediol; D006714:Homoserine; D005969:Glutamate Dehydrogenase
keywords: 1,3-Propanediol; Enzyme design; Glutamate dehydrogenase; Homoserine; Position-based prediction method; Protein engineering; Substrate specificity