Electrochemical investigations into kinase-catalyzed transformations of tau protein.

Abstract

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.

• Authors: Meghan K Rains, Sanela Martić, Daniel Freeman, Heinz Bernhard Kraatz
• Journal: ACS chemical neuroscience (ACS Chem Neurosci) Vol. 4 Issue 8 Pg. 1194-203 (Aug 21 2013) ISSN: 1948-7193 [Electronic] United States
• PMID: 23687953 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Ferrous Compounds
• MAS1 protein, human
• Metallocenes
• Proto-Oncogene Mas
• Proto-Oncogene Proteins
• tau Proteins
• Gold
• Adenosine Triphosphate
• tau-tubulin kinase
• Protein-Tyrosine Kinases
• Protein Serine-Threonine Kinases
• Glycogen Synthase Kinase 3
• ferrocene

Topics

• Adenosine Triphosphate (chemistry, metabolism)
• Electrochemical Techniques (methods)
• Ferrous Compounds (metabolism)
• Glycogen Synthase Kinase 3 (metabolism)
• Gold (chemistry)
• Humans
• Mass Spectrometry (methods)
• Metallocenes
• Phosphorylation
• Photoelectron Spectroscopy (methods)
• Protein Serine-Threonine Kinases (metabolism)
• Protein-Tyrosine Kinases (metabolism)
• Proto-Oncogene Mas
• Proto-Oncogene Proteins (metabolism)
• Surface Properties
• tau Proteins (chemistry, metabolism)