title: Single-molecule detection of phosphorylation-induced plasticity changes during ezrin activation.
abstract: Ezrin-radixin-moesin protein family provides a regulated link between the cortical actin cytoskeleton and the plasma membrane. Phosphorylation of ezrin has been functionally linked to membrane dynamics and plasticity. Our recent study demonstrated that phosphorylation of the conserved T567 residue of ezrin alters the physiology of gastric parietal cells. However, the molecular mechanism of phosphorylation-induced ezrin activation has remained elusive. Here we use atomic force microscopy (AFM) to probe phosphorylation-mediated activation of ezrin in single molecules. The phospho-mimicking and non-phosphorylatable mutant ezrin proteins were generated and purified to homogeneity. Comparative analyses of two ezrin mutants by AFM demonstrate the unfolding of the N- and C-terminal domains upon the phospho-activation. To measure the physical force underlying the inter-domain contact during mechanical unfolding, we probed the defined region of ezrin using the N-terminal ezrin coated onto the AFM tip. Comparative force measurements indicate that T567 phosphorylation-induced unfolding of ezrin favors the inter-molecular association. Taken together, these results provide molecular illustration of phosphorylation elicited functional activation of ERM proteins and indicate that stimulus-induced protein conformational change can be used as a signaling mechanism orchestrating cellular dynamics.
authors.affiliation: Division of Cellular Dynamics, Hefei National Laboratory for Physical Sciences, Hefei 230027, China.
authors.forename: Dan
authors.initials: D
authors.lastname: Liu
authors.forename: Ling
authors.initials: L
authors.lastname: Ge
authors.forename: Fengsong
authors.initials: F
authors.lastname: Wang
authors.forename: Hirohide
authors.initials: H
authors.lastname: Takahashi
authors.forename: Dongmei
authors.initials: D
authors.lastname: Wang
authors.forename: Zhen
authors.initials: Z
authors.lastname: Guo
authors.forename: Shige H
authors.initials: SH
authors.lastname: Yoshimura
authors.forename: Tarsha
authors.initials: T
authors.lastname: Ward
authors.forename: Xia
authors.initials: X
authors.lastname: Ding
authors.forename: Kunio
authors.initials: K
authors.lastname: Takeyasu
authors.forename: Xuebiao
authors.initials: X
authors.lastname: Yao
mesh_terms: D003598:Cytoskeletal Proteins / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D018625:Microscopy, Atomic Force; D010766:Phosphorylation; D017510:Protein Folding; D013912:Threonine / Q000235:genetics / Q000378:metabolism; D000099149:Ezrin
chemical_list: D003598:Cytoskeletal Proteins; D013912:Threonine; D000099149:Ezrin