Identification of Microtubule-Binding Domains on Microtubule-Associated Proteins by Major Coat Phage Display Technique

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• 作者：Binrui Cao ; Chuanbin Mao*
• 刊名：Biomacromolecules
• 出版年：2009
• 出版时间：March 9, 2009
• 年：2009
• 卷：10
• 期：3
• 页码：555-564
• 全文大小：403K
• 年卷期：v.10,no.3(March 9, 2009)
• ISSN：1526-4602

文摘

Microtubule is an important structural and functional component in cells. Microtubule-associated proteins (MAPs) are a class of proteins that can bind to microtubules and stabilize them to maintain their functions. However, not all the specific microtubule-binding domains on MAPs are clear. Here we report the study of microtubule-binding domains on MAPs from a new angle by biopanning a new type of phage-displayed random peptide library (called landscape phage library) against purified α- and β-tubulins. In the landscape phage library, billions of fd-tet phage clones are present and a unique 9-mer peptide is fused to each of the 3900 copies of major coat protein (pVIII) in each clone. The affinity-selected peptides derived from the biopanning were analyzed by the receptor ligand contacts (RELIC) suite of programs, which is a bioinformatics tool for combinatorial peptide analysis and identification of protein−ligand interaction sites. By using RELIC, 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. Our results confirm some known microtubule-binding domains and identify some new microtubule-binding domains and thus shed light into the mechanism of microtubule−MAPs interactions.