FPMOD: A Modeling Tool for Sampling the Conformational Space of Fusion Proteins Jason Chiang Department of Electrical and Computer Engineering Institute of Biomaterials and Biomedical Engineering June 9, 2006

MOTIVATION The existing modeling tools that can either create fusion proteins or sample conformational space, but none of them can do both tasks in a single platform. Once the conformational space is known, the average conformation for one fusion protein can then be predicted and so do their behaviors. Therefore, we developed a software called FPMOD (Fusion Protein MODeler). As a demonstration, FPMOD was used to predict the FRET efficiency changes of different FRET Ca2+ protein biosensors, which is one of the main interest of our lab. We verify the functionality of FPMOD by comparing the predicted simulation results to the in vitro experimental data.

Fluorescence Resonance Energy Transfer transfer of energy between donor and acceptor fluorescent proteins with partial spectral overlap Efficiency of FRET is distance- and orientation-dependent http://probes.invitrogen.com/handbook/boxes/0422.html

Simulation Process Fusion protein construction using script-lines Sample conformational space by rigid-body rotation

Conformational Space

Conformational Space

Conformational Space

Conformational Space

Conformational Space

RESULTS AND DISCUSSION So, we designed two FRET Ca2+ biosensor models, simulated their E% changes due to Ca2+-induction using FPMOD, and compared the simulation results to in vitro experimental data

Prediction vs. Experiment Donor emission peak Acceptor emission peak No Ca2+ with Ca2+ No Ca2+ With Ca2+ Change E% 17.0% 12.3% -4.7%

Prediction vs. Experiment No Ca2+ with Ca2+ No Ca2+ With Ca2+ Change E% 1.7% 3.1% 1.4%

CONCLUSIONS We developed a modeling tool called FPMOD to sample the possible conformations of fusion proteins using rigid-body domain rotation algorithms. FPMOD was then used to predict the E% changes of FRET Ca2+ biosensors and the results were qualitatively consistent with in vitro experimental data In the future, each fusion protein conformation will be quantitatively evaluated based on free energy, where lower energy is favored. Therefore, the sampled conformational space would account for protein forces such as van der Waals and electrostatic forces, as well.

ACKNOWLEDGEMENTS Supervisor: Dr. Kevin Truong Lab members: Isaac Li Elizabeth Pham National Science and Engineering Research Council (NSERC) Canadian Foundation of Innovation (CFI)

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