Algorithm development for modeling protein assemblies

Amyloid fibrils are associated with over 40 human diseases. For example, fibrils of human islet amyloid polypeptide (hIAPP), α-synuclein, and amyloid-β are pathological hallmarks of type 2 diabetes, Parkinson's disease and Alzheimer's disease, respectively. Understanding of fibril structure and that of the protofilaments that constitute the fibrils will help to reveal the mechanism of fibril formation in human diseases and to facilitate therapeutic intervention. However, the fibril structures of full-length amyloid proteins/peptides are difficult to determine by direct experimental approaches since they are neither soluble nor crystallizable. The goals of this thesis are to develop algorithms, programs and protocols for the modeling and determination of amyloid fibril structures based on experimental data mainly obtained from election paramagnetic resonance (EPR) spectroscopy and electron microscopy (EM), and then to use the resulting models for explaining previously unclear aspects of the experimental data and for aiding experimental design to obtain more structural details of the amyloid fibrils. In the thesis work, a simulation protocol for modeling amyloid protofilaments mainly based on EPR data was developed. This protocol was demonstrated to be able to produce structures of hIAPP protofilaments consistent with the experimental data. Then, to investigate the properties of the protofilament structures obtained and to generate fibril models consisting of more than one protofilament, a new program, MFIBRIL, was developed for flexible construction of fibrils from a monomeric unit. Several potential models of the hIAPP fibril were constructed using MFIBRIL and then refined by equilibration using molecular dynamics simulations in the NAMD software package. The refined models were evaluated by predicting the ring-to-ring distances using the MFIBRIL analysis tool and the inter-spin label distances and residue mobilities using PRONOX (another program developed in our laboratory) and comparing the predictions to the experimental data. This work led to identification of a favorable hIAPP fibril model consisting of two protofilaments with strong hydrophobic interactions between I26 and V32 surrounding hydrophilic interactions between S28 and T30 in the second β-strand (C-terminal) regions. The programs and protocols developed in this work are applicable to structural determination of other amyloid fibrils. In addition to the modeling of amyloid fibrils, two other projects are described in the thesis. The first is addition to the solvation function of WATGEN (a program developed in our laboratory for modeling the water network at protein-peptide interfaces) to protein-RNA interfaces and investigation of the features of solvated protein-RNA interfaces. The second project involves modeling of the transport mechanism of the human dipeptide transporter (hPEPT1).