Computational protocol: Characterization of Arabidopsis FPS Isozymes and FPS Gene Expression Analysis Provide Insight into the Biosynthesis of Isoprenoid Precursors in Seeds

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Protocol publication

[…] To construct homology models of Arabidopsis FPS1S and FPS2 that could be directly comparable with respect to their sequence differences, we chose as template the crystal structure of unliganded human FPS (PDB 2F7M) . The sequence identity of human FPS and FPS1S was 45% over 99% of its length (339 out of 343 amino acids) and between human FPS and FPS2 was 46% over 99% of its length (338 out of 342 amino acids). The phosphate ion and the four water molecules found in the active site of FPS in this structure were kept to maintain a stable conformation of the active site loops during modelling. First, we threaded the correct FPS1S and FPS2 sequences onto the template structure using Modeller 9.10 and then selected the 10 best models out of 1000 independent models on the basis of the Z-DOPE normalized scores, which ranged from –1.916 to –1.337 for FPS1S and from –1.949 to –1.529 for FPS2 (a Z-DOPE of less than –1 indicates a plausible model with 80% of the Cα atoms lying within 3.5 Å of their correct positions). Next, the top ten models for each protein were refined and minimized using the Rosetta force field and Monte Carlo sampling methods , and the improved models were clustered and analyzed to obtain a final model. The final models for FPS1S and FPS2 showed correct stereochemistry as assessed by MolProbity . […]

Pipeline specifications

Software tools MODELLER, MolProbity
Application Protein structure analysis
Organisms Arabidopsis thaliana, Escherichia coli