Studies on titin PEVK peptides and their interaction - Analytical Methods & Tools for Lyophilization

Studies on titin PEVK peptides and their interaction
Received 4 May 2006; revised 25 July 2006. Available online 15 August 2006.
Yingli Duana, 1, Joshua G. DeKeysera, 2, Srinivasan Damodaranb and Marion L. Greasera
Archives of Biochemistry and Biophysics
Volume 454, Issue 1 , 1 October 2006
ScienceDirect
Copyright ? 2006 Elsevier Inc. All rights reserved.
aMuscle Biology Laboratory, University of Wisconsin-Madison,1805 Linden Drive, Madison, WI 53706, USA
bDepartment of Food Science, University of Wisconsin-Madison, Madison, WI 53706, USA
Abstract
Experiments were conducted on several synthetic and expressed peptides from the PEVK region of titin, the giant muscle protein. Different secondary structure prediction methods based on amino acid sequence gave estimates ranging from over 70% a helical to no helix (totally disordered) for the polyE peptide corresponding to human exon 115. Circular dichroism (CD) experiments demonstrated that both the positively charged PPAK modules and the negatively charged PolyE repeats had similar spectral properties with disordered secondary structure predominating. Gel permeation chromatography showed that both PPAK and polyE peptides had 2?4 times larger Stokes radii than expected from their molecular mass. Mixtures of the oppositely charged titin peptides caused no change in apparent secondary structure as observed by circular dichroism or migration properties using native gel electrophoresis. Similarly addition of calcium did not alter the CD spectra or peptide electrophoretic mobility of the individual peptides or their mixtures. The properties of both the PPAK and polyE type peptides suggest that both had most of the characteristic properties to be classified as intrinsically disordered proteins.
Keywords: Titin; Muscle; Circular dichroism; Native electrophoresis; Ionic interaction; Secondary structure prediction; Intrinsically disordered proteins

Corresponding author. Fax: +1 608 265 3110.
1 Present address: University of Rochester, Rochester, NY 14627, USA.
2 Present address: The Pennsylvania University, University Park, PA 16802, USA.
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