Evolutionary Selection of Microtubule-Interacting Filamentous Phage with Thousands of Interaction Sites on the Side Walls
B. Cao, C. Mao
University of Oklahoma, US
Keywords: phage display, microtubule, tubulin
Abstract:Microtubule is an important structural and functional component in cells. The assembly and disassembly of microtubules are important for cell division. Thus microtubule-binding agents that can interfere with these processes are potential drugs that can inhibit mitotic progression. We hypothesize that microtubule-binding phage, which can be engineered to target cancer cells, can be a potential anti-cancer drug that can disrupt microtubule dynamics of cancer cells. We used landscape phage display technique to identify microtubule-binding phage by using purified alpha and beta tubulins as targets in biopanning. Because microtubule-associated proteins (MAPs) are a class of proteins that can bind to microtubules and stabilize them to maintain their functions, we compared affinity-selected tubulin-binding peptides with the MAPs by using the REceptor Ligand Contacts (RELIC) suite of programs, which is a bioinformatics tool for combinatorial peptide analysis and identification of protein-ligand interaction sites. The affinity-selected peptides were shown to have similarity with the sequences of two MAP families (MAP1 and MAP2/tau), thereby identifying putative microtubule-binding domains on these MAPs. The tubulin-binding affinity was also confirmed by using transmission electron microscopy (TEM) to characterize the interaction between affinity-selected tubulin-binding phage and tubulins.