1 research outputs found
Electrochemical Investigations into Kinase-Catalyzed Transformations of Tau Protein
The
formation of neurofibrillary tangles by hyperphosphorylated
tau is a well-recognized hallmark of Alzheimer’s disease. Resulting
from malfunctioning protein kinases, hyperphosphorylated tau is unable
to bind microtubules properly, causing it to self-associate and aggregate.
The effects of tau phosphorylation on tau conformation and aggregation
are still largely unexplored. The conformational analysis of tau and
its hyperphosphorylated forms is usually performed by a variety of
spectroscopic techniques, all of which require ample sample concentrations
and/or volumes. Here we report on the use of surface based electrochemical
techniques that allow for detection of conformational changes and
orientation of tau protein as a function of tau phosphorylation by
tyrosine and serine/threonine kinases. The electrochemical methods
utilize 5′-γ-ferrocenyl adenosine triphosphate (Fc-ATP)
derivative as a cosubstrate and tau immobilized on gold surface to
probe the role of the following protein kinases: Sarcoma related kinase
(Src), Abelson tyrosine kinase (Abl), tau-tubulin kinase (TTBK), proto-oncogene
tyrosine protein kinase Fyn (Fyn), and glycogen synthase kinase 3-β
(Gsk-3β). The single kinase and sequential kinase-catalyzed
Fc-phosphorylations modulate the electrochemical signal, pointing
to the dramatic changes around the Fc group in the Fc-phosphorylated
tau films. The location and orientation of the Fc-group in Fc-tau
film was investigated by the surface plasmon resonance based on antiferrocene
antibodies. Additional surface characterization of the Fc-tau films
by time-of-flight secondary ion-mass spectrometry and X-ray photoelectron
spectroscopy revealed that Fc-phosphorylations influence the tau orientation
and conformation on surfaces. When Fc-phosphorylations were performed
in solution, the subsequently immobilized Fc-tau exhibited similar
trends. This study illustrates the validity and the utility of the
labeled electrochemical approach for probing the changes in protein
film properties, conformation, and orientation as a function of the
enzymatically catalyzed modifications