title: Conformations within soluble oligomers and insoluble aggregates revealed by resonance energy transfer.
abstract: A fluorescently labeled 20-residue polyglutamic acid (polyE) peptide 20 amino acid length polyglutamic acid (E(20)) was used to study structural changes which occur in E(20) 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 E(20). 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 alpha-helical monomers to aggregated beta-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 E(20) 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.
authors.affiliation: Department of Chemistry, Oakland University, Rochester, MI 48309-4477, USA.
authors.forename: Jyothi L
authors.initials: JL
authors.lastname: Digambaranath
authors.forename: Loan
authors.initials: L
authors.lastname: Dang
authors.forename: Monika
authors.initials: M
authors.lastname: Dembinska
authors.forename: Andrew
authors.initials: A
authors.lastname: Vasyluk
authors.forename: John M
authors.initials: JM
authors.lastname: Finke
mesh_terms: D055598:Chemical Phenomena; D002942:Circular Dichroism; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D005456:Fluorescent Dyes / Q000737:chemistry; D010316:Particle Size; D011099:Polyglutamic Acid / Q000138:chemical synthesis / Q000737:chemistry*; D011485:Protein Binding; D011487:Protein Conformation; D012995:Solubility
chemical_list: D005456:Fluorescent Dyes; D011099:Polyglutamic Acid