(EMSL34)Parameter Development and Initial MD Simulation of Flavocytochrome Ifc3
EMSL Project ID
2388
Abstract
As part of the DOE Office of Science Program Notice 01-21 for Advanced Modeling and Simulation of Biological Systems, we have received funding for development of new modeling capabilities for the study of complex enzymatic reactions involved in the respiratory cycle of iron-reducing bacteria. The goal of this work is to provide the computational tools required for a detailed molecular-level characterization and understanding of the enzymatic reactivity of these bacteria through modeling and simulation, with a soluble flavocytochrome c3 of Shewanella frigidimarina as our target enzymatic system. Though the scope of the full project merits a grand challenge, we have broken up the initial research into several pilot projects that will provide guidance for a future grand challenge proposal. This pilot project involves developing the missing force field parameters for molecular dynamics simulation of S. frigidimarina flavocytochrome Ifc3 that contains four iron containing heme groups and flavin-adenine dinucleotide (FAD) plus the reactants, products and intermediates. We will expand the AMBER99 force field to include a consistent set of parameters for the heme groups, FAD and fumarate, using the recommended protocol, with geometries that we will determine from high-level (B3LYP/pVTZ) ab initio calculations and RESP-based partial atomic charges. We will develop the force field parameters for the porphyrin moiety with Fe in oxidation state III, as this is the appropriate state for the heme groups in the resting state of the enzyme. Flavins can exist in three different oxidation states (oxidized, semiquinone radical, and reduced) with multiple possible tautomeric forms for each oxidation state. X-ray crystal structures cannot be used to help deduce the oxidation state. Partial charges for the adenine moiety of FAD can be compared with the available parameters defined for DNA. Parameters will be developed for the two relevant isoalloxazine forms of FAD, the reduced anionic hydroquinone (activated) and the oxidized neutral isoalloxazine form. At physiological pH, the two carboxylic acid groups in the reactants and products (fumaric and succinic acid) are expected to be in the ionic state. We will determine partial atomic charges for the di-anionic, mono-protonated-anionic, and neutral forms. Torsion parameters will be adjusted to reproduce rotation barriers calculated from B3LYP/TZVP+ level theory. In addition, we will derive parameters for the partially reduced intermediate form of fumarate. Once all the parameters have been defined, we will carry out molecular dynamics simulations of the S. frigidimarina flavocytochrome Ifc3 in the open conformation starting with the 2.15 ? resolution crystal structure (PDB entry 1QO8) and in the closed form starting from the 1.80 ? resolution crystal structure (PDB entry 1QJD). We will insert fumarate into the active site of the closed conformation structure to replace the substrate analogue found in the crystal structure. Molecular dynamics simulations will be carried out for these two systems with FAD in the inactive and in the activated state. For these simulations the enzyme system will be solvated in water, using the SPC/E water model (Berendsen et al., 1987). To account for electrostatic interactions beyond the cutoff radius the particle-mesh Ewald method will be used. To complete such a large number of calculations will require the maximum 75,000 node-hours of computation time on an IBM-SMP like Jupiter or ECS1. Since one of our objectives is to determine the influence of the clamping domain motion on the structure of the active site and the conformation of the substrate in the active site, we expect to have to extend these molecular simulations for multiple nanoseconds. Such long simulations are beyond the scope of this pilot project.
Project Details
Project type
Capability Research
Start Date
2001-11-26
End Date
2002-09-30
Status
Closed
Released Data Link
Team
Principal Investigator
Team Members
Related Publications
Smith DMA, K Danyal, S Raugei, and LC Seefeldt. 2014. "Substrate Channel in Nitrogenase Revealed by a Molecular Dynamics Approach." Biochemistry 53(14):2278-2285. doi:10.1021/bi401313j