1. Conformations within soluble oligomers and insoluble aggregates revealed by resonance energy transfer
- Author
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Jyothi L. Digambaranath, Monika Dembinska, John M. Finke, Loan Dang, and Andrew Vasyluk
- Subjects
Circular dichroism ,Chemical Phenomena ,Chemistry ,Protein Conformation ,Circular Dichroism ,Organic Chemistry ,Polyglutamic acid ,Biophysics ,General Medicine ,Protein aggregation ,Biochemistry ,Oligomer ,Biomaterials ,chemistry.chemical_compound ,Crystallography ,Förster resonance energy transfer ,Protein structure ,Monomer ,Polyglutamic Acid ,Solubility ,Fluorescence Resonance Energy Transfer ,Protein folding ,Particle Size ,Fluorescent Dyes ,Protein Binding - Abstract
A fluorescently labeled 20-residue polyglutamic acid (polyE) peptide 20 amino acid length polyglutamic acid (E20) was used to study structural changes which occur in E20 as it co-aggregates with other unlabeled polyE peptides. Resonance energy transfer (RET) was performed using an o-aminobenzamide donor at the N-terminus and 3-nitrotyrosine acceptor at the C-terminus of E20. PolyE aggregates were not defined as amyloid, as they were nonfibrillar and did not bind congo red. Circular dichroism measurements indicate that polyE aggregation involves a transition from α-helical monomers to aggregated β-sheets. Soluble oligomers are also produced along with aggregates in the reaction, as determined through size exclusion chromatography. Time-resolved and steady-state RET measurements reveal four dominant E20 conformations: (1) a partially collapsed conformation (24 A donor–acceptor distance) in monomers, (2) an extended conformation in soluble oligomers (>29 A donor–acceptor distance), (3) a minor partially collapsed conformation (22 A donor-acceptor distance) in aggregates, and (4) a major highly collapsed conformation (13 A donor–acceptor distance) in aggregates. These findings demonstrate the use of RET as a means of determining angstrom-level structural details of soluble oligomer and aggregated states of proteins. © 2009 Wiley Periodicals, Inc. Biopolymers 93: 299–317, 2010. This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com
- Published
- 2009