TITLE:
Detection of cardiac myosin binding protein-C (cMyBP-C) by a phospho-specific PKD antibody in contracting rat cardiomyocytes
AUTHORS:
Ellen Dirkx, Freek G. Bouwman, Didier Vertommen, Edwin C. Mariman, Sakthivel Sadayappan, Jan F. C. Glatz, Joost J. F. P. Luiken, Guillaume J. van Eys
KEYWORDS:
Protein Kinase D; Cardiac Myosin Binding Protein-C; Phospho-Specific Antibody; Protein Phosphorylation
JOURNAL NAME:
Advances in Bioscience and Biotechnology,
Vol.4 No.4A,
April
29,
2013
ABSTRACT: Protein phosphorylation plays an important role in physiological
processes, such as muscle contraction. Phospho-specific antibodies have become powerful
tools to study these processes. Cardiac myosin binding protein-C (cMyBP-C) is
one of the proteins that make up the contractile apparatus of cardiomyocytes. Phosphorylation
of cMyBP-C is essential for normal cardiac function,
since dephosphorylation of this protein leads to its degradation and has
been associated with cardiomyopathy. One of the upstream kinases, which phosphorylate
cMyBP-C, is protein kinase D (PKD). While studying the role of PKD in cMyBP-C phosphorylation,
we tried to analyze phosphorylation of PKD with a phospho-specific PKD-Ser744/748
antibody. Contrary to the expected 115 kDa, a signal was found for a 150-kDa protein.
By MALDI-TOF mass spectrometry, we identified this protein to be cMyBP-C. These
data were confirmed by immunostaining using the p-PKD-Ser744/748 antibody, which
displayed a striated pattern similar to the one observed for a regular cMyBP-C antibody. To our knowledge there are no antibodies commercially available for phosphorylated cMyBP-C. Thus, the p-PKD-Ser744/748 antibody can accelerate research into the
role of cMyBP-C phosphorylation in cardiomyocytes.