(386b) In Silico Protein Fragmentation Reveals The Importance Of Critical Nuclei In Domain Reassembly

Protein complementation assays (PCAs) based on split protein fragments have become powerful research tools that facilitate the study and engineering of intracellular protein-protein interactions. These assays are based on the observation that: (1) a given protein can be split into individual fragments that by themselves remain inactive; and (2) the fragments can reassemble under the appropriate cellular conditions to yield the original properly folded and active structure. However, one of the experimental limitations of such systems is the observation that the folding of a protein from its fragments is dramatically slower relative to the folding of the parent (unsplit) protein. This is due in large part to our poor understanding of how certain factors (i.e. location of split site in primary sequence, size of protein fragments, etc.) contribute to the efficient reconstitution of the folded protein. To address some of the theoretical factors that contribute to the thermodynamics of protein reassembly, we have used a minimalist on-lattice model to study how the position where a protein is split contributes to the efficiency of its folding. In particular, we have analyzed how the reassembly process is affected by the disruption of the ?folding nucleus,? which is a subset of residues that is critical to the formation of the transition state that leads to productive folding. We will discuss how our results have elucidated some of the key mechanisms underlying the folding of protein fragments and how this insight might potentially be used to rationally design optimized split-protein systems for different biomolecular applications.