Conformations within soluble oligomers and insoluble aggregates revealed by resonance energy transfer

Digambaranath, Jyothi L.; Dang, Loan; Dembinska, Monika; Vasyluk, Andrew; Finke, John M.

doi:10.1002/bip.21324

Cited by 6 publications

(16 citation statements)

References 65 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The CD spectrum is highly similar to that of protein aggregates characterized elsewhere (Digambaranath et al, 2010). ( C ) Overlay of SEC-MALS chromatograms of SAS-5 FL samples at multiple concentrations, showing scaled light scattering intensity vs elution volume.…”

Section: Resultssupporting

confidence: 65%

The Caenorhabditis elegans protein SAS-5 forms large oligomeric assemblies critical for centriole formation

Rogala

Dynes

Hatzopoulos

et al. 2015

eLife

View full text Add to dashboard Cite

Centrioles are microtubule-based organelles crucial for cell division, sensing and motility. In Caenorhabditis elegans, the onset of centriole formation requires notably the proteins SAS-5 and SAS-6, which have functional equivalents across eukaryotic evolution. Whereas the molecular architecture of SAS-6 and its role in initiating centriole formation are well understood, the mechanisms by which SAS-5 and its relatives function is unclear. Here, we combine biophysical and structural analysis to uncover the architecture of SAS-5 and examine its functional implications in vivo. Our work reveals that two distinct self-associating domains are necessary to form higher-order oligomers of SAS-5: a trimeric coiled coil and a novel globular dimeric Implico domain. Disruption of either domain leads to centriole duplication failure in worm embryos, indicating that large SAS-5 assemblies are necessary for function in vivo.DOI: http://dx.doi.org/10.7554/eLife.07410.001

show abstract

Section: Resultssupporting

confidence: 65%

The Caenorhabditis elegans protein SAS-5 forms large oligomeric assemblies critical for centriole formation

Rogala

Dynes

Hatzopoulos

et al. 2015

eLife

View full text Add to dashboard Cite

show abstract

“…Samples from both the proteins obtained after different incubation times were spun down at 12,000 rpm at 37°C for 10 min to separate the soluble protein fraction (supernatant) from the insoluble fraction (pellet) [42]. Concentrations of the soluble protein fractions were determined by measuring the absorbance at 280 nm using known values of the extinction co-efficients of both proteins.…”

Section: Resultsmentioning

confidence: 99%

Characterization of the Oligomerization and Aggregation of Human Serum Amyloid A

et al. 2013

View full text Add to dashboard Cite

The fibrillation of Serum Amyloid A (SAA) – a major acute phase protein – is believed to play a role in the disease Amyloid A (AA) Amyloidosis. To better understand the amyloid formation pathway of SAA, we characterized the oligomerization, misfolding, and aggregation of a disease-associated isoform of human SAA – human SAA1.1 (hSAA1.1) – using techniques ranging from circular dichroism spectroscopy to atomic force microscopy, fluorescence spectroscopy, immunoblot studies, solubility measurements, and seeding experiments. We found that hSAA1.1 formed alpha helix-rich, marginally stable oligomers in vitro on refolding and cross-beta-rich aggregates following incubation at 37°C. Strikingly, while hSAA1.1 was not highly amyloidogenic in vitro, the addition of a single N-terminal methionine residue significantly enhanced the fibrillation propensity of hSAA1.1 and modulated its fibrillation pathway. A deeper understanding of the oligomerization and fibrillation pathway of hSAA1.1 may help elucidate its pathological role.

show abstract

“…Two peptides were synthesized for this study: (1) ABZ‐K 2 Q 16 K 2 ‐Y (DQ 16 ) and (2) ABZ‐K 2 Q 16 K 2 ‐YNO 2 (DQ 16 A). In these peptides, ABZ denotes an n‐terminal o ‐aminobenzamide (ABZ; FRET donor), Y denotes a c‐terminal tyrosine, and YNO 2 denotes a c‐terminal 3‐nitrotyrosine (FRET acceptor) 49, 57, 58, 68. The peptides were obtained through custom synthesis (Anaspec) with purity >95% was achieved with HPLC purification and confirmed through MALDI mass spectrometry and molecular weight analysis on a Beckman XLA analytical centrifuge.…”

Section: Methodsmentioning

confidence: 99%

“…FRET has been an invaluable approach to study the structural ensemble of unfolded proteins and intrinsically disordered proteins in prior studies 19, 46–66. The present FRET study is modeled after previous successful FRET studies of polyglutamic acid helix‐coil transitions and aggregation 57, 58.…”

Section: Introductionmentioning

confidence: 99%

“…The present FRET study is modeled after previous successful FRET studies of polyglutamic acid helix‐coil transitions and aggregation 57, 58. As in these previous experiments, the present study used both steady‐state and time‐resolved fluorescence to determine the end‐to‐end distance of the canonical polyglutamine peptide KKQ 16 KK with an aminobenzamide donor at the n‐terminus and nitrotyrosinamide at the c‐terminus (DQ 16 A) 57, 58. This end‐to‐end distance will be used, along with previously published donor–acceptor distances determined by FRET, triplet‐state quenching rates, and radius of gyration scaling to assess the accuracy of different theoretical models 19, 25–27.…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

An accurate model of polyglutamine

et al. 2011

Self Cite

View full text Add to dashboard Cite

Polyglutamine repeats in proteins are highly correlated with amyloid formation and neurological disease. To better understand the molecular basis of glutamine repeat diseases, structural analysis of polyglutamine peptides as soluble monomers, oligomers, and insoluble amyloid fibrils is necessary. In this study, fluorescence resonance energy transfer (FRET) experiments and molecular dynamics simulations using different theoretical models of polyglutamine were conducted. This study demonstrates that a previously proposed simple C(α)C(β) model of polyglutamine, denoted as FCO, accurately reproduced the present FRET results and the results of previously published FRET, triplet-state quenching, and fluorescence correlation studies. Other simple C(α)C(β) models with random coil and extended β-strand parameters, and all-atom models with parm96 and parm99SB force fields, did not match the FRET result well. The FCO is an intrinsically disordered model with a high-effective persistence length producing extended peptides at short lengths (Q(N) < 10). Because of an increasing number of attractive Q-Q interactions at longer lengths, the FCO model becomes increasingly more compact at lengths between Q(N) ∼ 10-16 and is as compact as many folded proteins at Q(N) > 16.

show abstract

Conformations within soluble oligomers and insoluble aggregates revealed by resonance energy transfer

Cited by 6 publications

References 65 publications

The Caenorhabditis elegans protein SAS-5 forms large oligomeric assemblies critical for centriole formation

The Caenorhabditis elegans protein SAS-5 forms large oligomeric assemblies critical for centriole formation

Characterization of the Oligomerization and Aggregation of Human Serum Amyloid A

An accurate model of polyglutamine

Contact Info

Product

Resources

About