Computational protocol: Structural and functional definition of the specificity of a novel caspase-3 inhibitor, Ac-DNLD-CHO

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Protocol publication

[…] Molecular visualization was carried out in a DS ViewerPro (Accelrys, Inc., San Diego, CA). The coordinates of caspases-3, -7, -8, and -9 were obtained from the Protein Data Bank (PDB) (codes 1PAU, 1F1J, 1F9E, and 1JXQ). Water was removed from the PDB files. The crystal structures of caspases-3, -7, -8, and -9 include coordinates of Ac-DEVD-CHO, Ac-DEVD-CHO, Z-DEVD-CHO, and Glu-Val-Dehydroxymethylaspartic acid, respectively, as inhibitors. Since it is known that Ac-DEVD-CHO potently inhibits the activities of caspases [], we constructed the models of the Ac-DEVD-CHO/caspase complex. To construct the complex structure of caspase-8/Ac-DEVD-CHO, the benzyloxy-carbonyl group of Z-DEVD-CHO was displaced by the acetyl group on the DS ViewerPro. The complex structure of Ac-DEVD-CHO/caspase-9 was constructed by superposition with that of the Ac-DEVD-CHO/caspase-8. The superposition was performed using the McLaghlan algorithm [] as implemented in the ProFit program []. The geometries of these complex structures were subsequently optimized using the GROMACS program [,]. The coordinate of Ac-DEVD-CHO was converted to a GROMACS topology file by the Dundee PRODRG2 Server []. Energy minimizations of the complex structures using a GROMACS forcefield were performed with the steepest descent algorithm. [...] Molecular docking was carried out using AutoDock3.0 [] on a COMPAQ Alphastation DS20E (double 833 MHz processors and 1024 MB of memory). The binding free energy scoring function in the AutoDock is based on an empirical function derived by linear regression analysis of a large set of diverse protein-ligand complexes with known inhibition constants. There are many successful examples of structures of protein-ligand systems studied by the AutoDock program [,]. The docking energy grid (grid maps with 60 × 60 × 60 points, grid spacing 0.375 Å) was produced with the AutoGrid program []. The inhibitor centers in the complex structures were positioned at the grid center. The Lamarckian Genetic Algorithm was utilized, and energy evaluations were set at 3 × 106. Each simulation was performed a total of 20 times. Other parameters were default values. The initial conformations of caspase-3 inhibitors, Ac-DNLD-CHO, Ac-DEVD-CHO, Ac-DQTD-CHO, and Ac-DMQD-CHO, were built using coordinates that replaced the side chains of the Ac-DEVD-CHO in the complex structures with the side chains of the inhibitors. Rotational bonds in the inhibitors were assigned with the program AutoTors []. All torsions except the peptide bonds were unconstrained during the docking. Based on the docking data, the lowest-energy docking mode was used to analyze the potency and selectivity of the caspase-3 peptide inhibitors. […]

Pipeline specifications

Software tools GROMACS, PRODRG, AutoDock
Applications Drug design, Protein interaction analysis
Organisms Dipturus trachyderma
Diseases Virus Diseases, Neurodegenerative Diseases, Mitochondrial Diseases