Publications

Cait MacPhee

Edinburgh Research Explorer

Recent papers

Interfacial self-assembly of a bacterial hydrophobin

KM Bromley, RJ Morris,L Hobley, GB Brandani, RMC Gillespie, M McCluskey, U Zachariae, D Marenduzzo, NR Stanley-Wall, and CE MacPhee

Proc Nat'l Acad Sci USA 112: 5419-5424 (2015)

In the natural environment the majority of bacteria live within the confines of a structured social community called a biofilm. The stability of biofilms arises from the extracellular matrix, which consists of proteins, polysaccharides, and extracellular DNA. One of these proteins, BslA, forms a hydrophobic “raincoat” at the surface of the biofilm. We have uncovered the mechanism that enables this protein to function, revealing a structural metamorphosis from a form that is stable in water to a structure that prefers the interface where it self-assembles with nanometer precision to form a robust film. Our findings have wide-ranging implications, from the disruption of harmful bacterial biofilms to the generation of nanoscale materials.

doi: 10.1073/pnas.1419016112

A kinetic study of ovalbumin fibril formation: the importance of fragmentation and end-joining

JMD Kalapothakis, RJ Morris, J Szavits-Nossan, K Eden, S Covill, S Tabor, J Gillam, PE Barran, RJ Allen and CE MacPhee

Biophys J 108: 2300-2311 (2015)

The ability to control the morphologies of biomolecular aggregates is a central objective in the study of self-assembly processes. The development of predictive models offers the surest route for gaining such control. Under the right conditions, proteins will self-assemble into fibers that may rearrange themselves even further to form diverse structures, including the formation of closed loops. In this study, chicken egg white ovalbumin is used as a model for the study of fibril loops. By monitoring the kinetics of self-assembly, we demonstrate that loop formation is a consequence of end-to-end association between protein fibrils. A model of fibril formation kinetics, including end-joining, is developed and solved, showing that end-joining has a distinct effect on the growth of fibrillar mass density (which can be measured experimentally), establishing a link between self-assembly kinetics and the underlying growth mechanism. These results will enable experimentalists to infer fibrillar morphologies from an appropriate analysis of self-assembly kinetic data.

doi:10.1016/j.bpj.2015.03.021

Giving structure to the biofilm matrix: an overview of individual strategies and emerging common themes

L Hobley, C Harkins, CE MacPhee and NR Stanley-Wall

FEMS Microbiol Rev fuv015 (2015)

Biofilms are communities of microbial cells that underpin diverse processes including sewage bioremediation, plant growth promotion, chronic infections and industrial biofouling. The cells resident in the biofilm are encased within a self-produced exopolymeric matrix that commonly comprises lipids, proteins that frequently exhibit amyloid-like properties, eDNA and exopolysaccharides. This matrix fulfils a variety of functions for the community, from providing structural rigidity and protection from the external environment to controlling gene regulation and nutrient adsorption. Critical to the development of novel strategies to control biofilm infections, or the capability to capitalize on the power of biofilm formation for industrial and biotechnological uses, is an in-depth knowledge of the biofilm matrix. This is with respect to the structure of the individual components, the nature of the interactions between the molecules and the three-dimensional spatial organization. We highlight recent advances in the understanding of the structural and functional role that carbohydrates and proteins play within the biofilm matrix to provide three-dimensional architectural integrity and functionality to the biofilm community. We highlight, where relevant, experimental techniques that are allowing the boundaries of our understanding of the biofilm matrix to be extended using Escherichia coli, Staphylococcus aureus, Vibrio cholerae, and Bacillus subtilis as exemplars.

DOI: http://dx.doi.org/10.1093/femsre/fuv015

Membrainy: a 'smart', unified membrane analysis tool

M Carr, CE MacPhee

Source Code for Biology and Medicine 10: 3 (2015)

The study of biological membranes using Molecular Dynamics has become an increasingly popular means by which to investigate the interactions of proteins, peptides and potentials with lipid bilayers. These interactions often result in changes to the properties of the lipids which can modify the behaviour of the membrane. Membrainy is a unified membrane analysis tool that contains a broad spectrum of analytical techniques to enable: measurement of acyl chain order parameters; presentation of 2D surface and thickness maps; determination of lateral and axial headgroup orientations; measurement of bilayer and leaflet thickness; analysis of the annular shell surrounding membrane-embedded objects; quantification of gel percentage; time evolution of the transmembrane voltage; area per lipid calculations; and quantification of lipid mixing/demixing entropy. Results Each analytical component within Membrainy has been tested on a variety of lipid bilayer systems and was found to be either comparable to or an improvement upon existing software. For the analytical techniques that have no direct comparable software, our results were confirmed with experimental data. Membrainy is a user-friendly, intelligent membrane analysis tool that automatically interprets a variety of input formats and force fields, is compatible with both single and double bilayers, and capable of handling asymmetric bilayers and lipid flip-flopping. Membrainy has been designed for ease of use, requiring no installation or configuration and minimal user-input to operate.

doi:10.1186/s13029-015-0033-7
Find membrainy here

Competition between Primary Nucleation and Autocatalysis in Amyloid Fibril Self-Assembly

K Eden, R Morris, J Gillam, CE MacPhee, RJ Allen

Biophys J 108, 632-643 (2015)

Kinetic measurements of the self-assembly of proteins into amyloid fibrils are often used to make inferences about molecular mechanisms. In particular, the lag time - the quiescent period before aggregates are detected - is often found to scale with the protein concentration as a power law, whose exponent has been used to infer the presence or absence of autocatalytic growth processes such as fibril fragmentation. Here we show that experimental data for lag time versus protein concentration can show signs of kinks: clear changes in scaling exponent, indicating changes in the dominant molecular mechanism determining the lag time. Classical models for the kinetics of fibril assembly suggest that at least two mechanisms are at play during the lag time: primary nucleation and autocatalytic growth. Using computer simulations and theoretical calculations, we investigate whether the competition between these two processes can account for the kinks which we observe in our and others' experimental data. We derive theoretical conditions for the crossover between nucleation-dominated and growth-dominated regimes, and analyze their dependence on system volume and autocatalysis mechanism. Comparing these predictions to the data, we find that the experimentally observed kinks cannot be explained by a simple crossover between nucleation-dominated and autocatalytic growth regimes. Our results show that existing kinetic models fail to explain detailed features of lag time versus concentration curves, suggesting that new mechanistic understanding is needed. More broadly, our work demonstrates that care is needed in interpreting lag-time scaling exponents from protein assembly data.

doi:10.1016/j.bpj.2014.11.3465

Relating gas phase to solution conformations: lessons from disordered proteins

R Beveridge, AS Phillips, L Denbigh, HM Saleem, CE MacPhee, PE Barran

Proteomics (2015)

In recent years both mass spectrometry (MS) and ion mobility mass spectrometry (IM-MS) have been developed as techniques with which to study proteins that lack a fixed tertiary structure but may contain regions of that form secondary structure elements transiently, namely intrinsically disordered proteins (IDPs). IM-MS is a suitable method for the study of IDPs which provides an insight to conformations that are present in solution, potentially enabling the analysis of lowly populated structural forms. Here we describe the IM-MS data of two IDPs; α-Synuclein (α-Syn) which is implicated in Parkinson's Disease, and Apolipoprotein C-II (ApoC-II) which is involved in cardiovascular diseases. We report an apparent discrepancy in the way that ApoC-II behaves in the gas phase. While most IDPs, including α-Syn, present in many charge states and a wide range of rotationally averaged collision cross sections (CCSs), ApoC-II presents in just 4 charge states and a very narrow range of CCSs, independent of solution conditions. Here, we compare MS and IM-MS data of both proteins, and rationalise the differences between the proteins in terms of different ionisation processes which they may adhere to.

DOI: 10.1002/pmic.201400605

Electron capture dissociation and drift tube ion mobility-mass spectrometry coupled with site directed mutations provide insights into the conformational diversity of a metamorphic protein

SR Harvey, M Porrini, R Tyler, C MacPhee, B Volkman, P Barran

Phys Chem Chem Phys 17: 10538-10550 (2015)

Ion mobility mass spectrometry can be combined with data from top-down sequencing to discern adopted conformations of proteins in the absence of solvent. This multi-technique approach has particular applicability for conformationally dynamic systems. Previously, we demonstrated the use of drift tube ion mobility-mass spectrometry (DT IM-MS) and electron capture dissociation (ECD) to study the metamorphic protein lymphotactin (Ltn). Ltn exists in equilibrium between distinct monomeric (Ltn10) and dimeric (Ltn40) folds, both of which can be preserved and probed in the gas-phase. Here, we further test this mass spectrometric framework, by examining two site directed mutants of Ltn, designed to stabilise either distinct fold in solution, in addition to a truncated form consisting of a minimum model of structure for Ltn10. The truncated mutant has similar collision cross sections to the wild type (WT), for low charge states, and is resistant to ECD fragmentation. The monomer mutant (CC3) presents in similar conformational families as observed previously for the WT Ltn monomer. As with the WT, the CC3 mutant is resistant to ECD fragmentation at low charge states. The dimer mutant W55D is found here to exist as both a monomer and dimer. As a monomer W55D exhibits similar behaviour to the WT, but as a dimer presents a much larger charge state and collision cross section range than the WT dimer, suggesting a smaller interaction interface. In addition, ECD on the W55D mutant yields greater fragmentation than for the WT, suggesting a less stable β-sheet core. The results highlight the power of MS to provide insight into dynamic proteins, providing further information on each distinct fold of Ltn. In addition we observe differences in the fold stability following single or double point mutations. This approach, therefore, has potential to be a useful tool to screen for the structural effects of mutagenesis, even when sample is limited.

DOI: 10.1039/C4CP05136J

Conformational dynamics of α-synuclein: insights from mass spectrometry

AS Phillips, AF Gomes, JMD Kalapothakis, JE Gillam, J Gasparavicius, FC Gotto, T Kunath, CE MacPhee, PE Barran

Analyst 140: 3070-3081 (2015)

The aggregation and deposition of α-synuclein in Lewy bodies is associated with the progression of Parkinson's disease. Here, Mass Spectrometry (MS) is used in combination with Ion Mobility (IM), chemical crosslinking and Electron Capture Dissociation (ECD) to probe transient structural elements of α-synuclein and its oligomers. Each of these reveals different aspects of the conformational heterogeneity of this 14 kDa protein. IM-MS analysis indicates that this protein is highly disordered, presenting in positive ionisation mode with a charge state range of 5 ≤ z ≤ 21 for the monomer, along with a collision cross section range of ∼1600 Å2. Chemical crosslinking applied in conjunction with IM-MS captures solution phase conformational families enabling comparison with those exhibited in the gas phase. Crosslinking IM-MS identifies 3 distinct conformational families, Compact (∼1200 Å2), Extended (∼1500 Å2) and Unfolded (∼2350 Å2) which correlate with those observed in solution. ECD-Fourier Transform-Ion Cyclotron Resonance Mass Spectrometry (ECD-FT-ICR MS) highlights the effect of pH on α-synuclein structure, identifying the conformational flexibility of the N and C termini as well as providing evidence for structure in the core and at times the C terminus. A hypothesis is proposed for the variability displayed in the structural rearrangement of α-synuclein following changes in solution pH. Following a 120 h aggregation time course, we observe an increase in the ratio of dimer to monomer, but no gross conformational changes in either, beyond the significant variations that are observed day-to-day from this conformationally dynamic protein.

DOI: 10.1039/C4AN02306D

A Mass-Spectrometry-Based Framework To Define the Extent of Disorder in Proteins

R Beveridge, S Covill, K J Pacholarz, JMD Kalapothakis, CE MacPhee, P E Barran

Anal Chem 86: 10979-10991 (2014)

In the past decade, mass spectrometry (MS) coupled with electrospray ionization (ESI) has been extensively applied to the study of intact proteins and their complexes, often without the requirement of labels. Solvent conditions (for example, pH, ionic strength, and concentration) affect the observed desolvated species; the ease of altering such extrinsic factors renders ESI-MS an appropriate method by which to consider the range of conformational states that proteins may occupy, including natively folded, disordered and amyloid. Rotationally averaged collision cross sections of the ionized forms of proteins, provided by the combination of mass spectrometry and ion mobility (IM-MS), are also instructive in exploring conformational landscapes in the absence of solvent. Here, we ask the following question: "If the only technique you had was ESI-IM-MS, what information would it provide on the structural preferences of an unknown protein?" We have selected 20 different proteins, both monomeric and multimeric, ranging in mass from 2846 Da (melittin) to 150 kDa (Immunoglobulin G), and we consider how they are presented to a mass spectrometer under different solvent conditions. Mass spectrometery allows us to distinguish which of these proteins are structured (melittin, human beta defensin 1, truncated human lymphotactin, Cytochrome C, holo hemoglobin-α, ovalbumin, human transthyretin, avidin, bovine serum albumin, concanavalin, human serum amyloid protein, and Immunoglobulin G) from those that contain at least some regions of disorder (human lymphotactin, N-terminal p53, α-Synuclein, N-terminal MDM2, and p53 DNA binding domain) or denatured due to solvent conditions (ubiquitin, apo hemoglobin-α, apo hemoglobin-β) by considering two experimental parameters: the range of charge states occupied by the protein (Δz) and the range of collision cross sections in which the protein is observed (ΔCCS). We also provide a simple model to predict the difference between the collision cross sections of the most compact and the most extended form of a given protein, based on the volume of the amino acids it contains. We compare these calculated parameters with experimental values. In addition, we consider the occupancy of conformations based on the intensities of ions in the mass spectra. This allows us to qualitatively predict the potential energy landscape of each protein. Our empirical approach to assess order or disorder is shown to be more accurate than the use of charge hydropathy plots, which are frequently used to predict disorder, and could provide an initial route to characterization. Finally, we present an ESI-IM-MS methodology to determine if a given protein is structured or disordered.

DOI: 10.1021/ac5027435

Dissecting the Dynamic Conformations of the Metamorphic Protein Lymphotactin

SR Harvey, M Porrini, A Konijnenberg, DJ Clarke, RC Tyler, PRR Langridge-Smith, CE MacPhee, BF Volkman, PE Barran

Barran J Phys Chem B 118: 12348-12359 (2014)

A mass spectrometer provides an ideal laboratory to probe the structure and stability of isolated protein ions. Interrogation of each discrete mass/charge-separated species enables the determination of the intrinsic stability of a protein fold, gaining snapshots of unfolding pathways. In solution, the metamorphic protein lymphotactin (Ltn) exists in equilibrium between two distinct conformations, a monomeric (Ltn10) and a dimeric (Ltn40) fold. Here, we use electron capture dissociation (ECD) and drift tube ion mobility-mass spectrometry (DT IM-MS) to analyze both forms and use molecular dynamics (MD) to consider how the solution fold alters in a solvent-free environment. DT IM-MS reveals significant conformational flexibility for the monomer, while the dimer appears more conformationally restricted. These findings are supported by MD calculations, which reveal how salt bridges stabilize the conformers in vacuo. Following ECD experiments, a distinctive fragmentation pattern is obtained for both the monomer and dimer. Monomer fragmentation becomes more pronounced with increasing charge state especially in the disordered regions and C-terminal α-helix in the solution fold. Lower levels of fragmentation are seen in the β-sheet regions and in regions that contain salt bridges, identified by MD simulations. The lowest charge state of the dimer for which we obtain ECD data ([D+9H]9+) exhibits extensive fragmentation with no relationship to the solution fold and has a smaller collision cross section (CCS) than charge states 10-13+, suggesting a "collapsed" encounter complex. Other charge states of the dimer, as for the monomer, are resistant to fragmentation in regions of β-sheets in the solution fold. This study provides evidence for preservation and loss of global fold and secondary structural elements, providing a tantalizing glimpse into the power of the emerging field of native top-down mass spectrometry.

DOI: 10.1021/jp504997k

Inherent variability in the kinetics of autocatalytic protein self-assembly

J Szavits-Nossan, K Eden, RJ Morris, CE MacPhee, MR Evans, RJ Allen

Phys Rev Lett 113: 098101 (2014)

In small volumes, the kinetics of filamentous protein self-assembly is expected to show significant variability, arising from intrinsic molecular noise. This is not accounted for in existing deterministic models. We introduce a simple stochastic model including nucleation and autocatalytic growth via elongation and fragmentation, which allows us to predict the effects of molecular noise on the kinetics of autocatalytic self-assembly. We derive an analytic expression for the lag-time distribution, which agrees well with experimental results for the fibrillation of bovine insulin. Our expression decomposes the lag-time variability into contributions from primary nucleation and autocatalytic growth and reveals how each of these scales with the key kinetic parameters. Our analysis shows that significant lag-time variability can arise from both primary nucleation and from autocatalytic growth and should provide a way to extract mechanistic information on early-stage aggregation from small-volume experiments.

DOI: 10.1103/PhysRevLett.113.098101

Quantifying disorder through conditional entropy: an application to fluid mixing

G Brandani, M Schor, CE MacPhee, U Zachariae, D Marenduzzo

PLoS one 8: e65617 (2013)

In this paper, we present a method to quantify the extent of disorder in a system by using conditional entropies. Our approach is especially useful when other global, or mean field, measures of disorder fail. The method is equally suited for both continuum and lattice models, and it can be made rigorous for the latter. We apply it to mixing and demixing in multicomponent fluid membranes, and show that it has advantages over previous measures based on Shannon entropies, such as a much diminished dependence on binning and the ability to capture local correlations. Further potential applications are very diverse, and could include the study of local and global order in fluid mixtures, liquid crystals, magnetic materials, and particularly biomolecular systems.

DOI: 10.1371/journal.pone.0065617

Mechanistic and environmental control of the prevalence and lifetime of amyloid oligomers

R Morris, K Eden, R Yarwood, RJ Allen & CE MacPhee

Nature Comm 4: 1891 (2013)

Amyloid fibrils are self-assembled protein aggregates implicated in a number of human diseases. Fragmentation-dominated models for the self-assembly of amyloid fibrils have had important successes in explaining the kinetics of amyloid fibril formation but predict fibril length distributions that do not match experiments. Here we resolve this inconsistency using a combination of experimental kinetic measurements and computer simulations. We provide evidence for a structural transition that occurs at a critical fibril mass concentration, or CFC, above which fragmentation of fibrils is suppressed. Our simulations predict the formation of distinct fibril length distributions above and below the CFC, which we confirm by electron microscopy. These results point to a new picture of amyloid fibril growth in which structural transitions that occur during self-assembly have strong effects on the final population of aggregate species with small, and potentially cytotoxic, oligomers dominating for long periods of time at protein concentrations below the CFC.

doi:10.1038/ncomms2909

Modelling amyloid fibril formation kinetics: mechanisms of nucleation and growth

JE Gillam & CE MacPhee

J Phys Condens Matt 25: 373101 (2013)

Amyloid and amyloid-like fibrils are self-assembling protein nanostructures, of interest for their robust material properties and inherent biological compatibility as well as their putative role in a number of debilitating mammalian disorders. Understanding fibril formation is essential to the development of strategies to control, manipulate or prevent fibril growth. As such, this area of research has attracted significant attention over the last half century. This review describes a number of different models that have been formulated to describe the kinetics of fibril assembly. We describe the macroscopic implications of mechanisms in which secondary processes such as secondary nucleation, fragmentation or branching dominate the assembly pathway, compared to mechanisms dominated by the influence of primary nucleation. We further describe how experimental data can be analysed with respect to the predictions of kinetic models.Amyloid and amyloid-like fibrils are self-assembling protein nanostructures, of interest for their robust material properties and inherent biological compatibility as well as their putative role in a number of debilitating mammalian disorders. Understanding fibril formation is essential to the development of strategies to control, manipulate or prevent fibril growth. As such, this area of research has attracted significant attention over the last half century. This review describes a number of different models that have been formulated to describe the kinetics of fibril assembly. We describe the macroscopic implications of mechanisms in which secondary processes such as secondary nucleation, fragmentation or branching dominate the assembly pathway, compared to mechanisms dominated by the influence of primary nucleation. We further describe how experimental data can be analysed with respect to the predictions of kinetic models.

doi:10.1088/0953-8984/25/37/373101

BslA is a self-assembling bacterial hydrophobin that coats the Bacillus subtilis biofilm

L Hobley, A Ostrowski, FV Rao, KM Bromley, MPorter, AR Prescott, CE MacPhee, DMF Van Aalten, NR Stanley-Wall

Proc Nat'l Acad Sci USA 110: 13600-13605 (2013)

Biofilms represent the predominant mode of microbial growth in the natural environment. Bacillus subtilis is a ubiquitous Gram-positive soil bacterium that functions as an effective plant growth-promoting agent. The biofilm matrix is composed of an exopolysaccharide and an amyloid fiber-forming protein, TasA, and assembles with the aid of a small secreted protein, BslA. Here we show that natively synthesized and secreted BslA forms surface layers around the biofilm. Biophysical analysis demonstrates that BslA can self-assemble at interfaces, forming an elastic film. Molecular function is revealed from analysis of the crystal structure of BslA, which consists of an Ig-type fold with the addition of an unusual, extremely hydrophobic "cap" region. A combination of in vivo biofilm formation and in vitro biophysical analysis demonstrates that the central hydrophobic residues of the cap are essential to allow a hydrophobic, nonwetting biofilm to form as they control the surface activity of the BslA protein. The hydrophobic cap exhibits physiochemical properties remarkably similar to the hydrophobic surface found in fungal hydrophobins; thus, BslA is a structurally defined bacterial hydrophobin. We suggest that biofilms formed by other species of bacteria may have evolved similar mechanisms to provide protection to the resident bacterial community.

doi: 10.1073/pnas.1306390110

Mass spectrometry methods for intrinsically disordered proteins

R Beveridge, Q Chappuis, CE MacPhee, PE Barran

Analyst 138: 32-42 (2013)

In the last ten years mass spectrometry has emerged as a powerful biophysical technique capable of providing unique insights into the structure and dynamics of proteins. Part of this explosion in use involves investigations of the most recently 'discovered' subset of proteins: the so-called 'Intrinsically Disordered' or 'Natively Unstructured' proteins. A key advantage of the use of mass spectrometry to study intrinsically disordered proteins (IDPs) is its ability to test biophysical assertions made about why they differ from structured proteins. For example, from the charge state distribution presented by a protein following nano-electrospray (n-ESI) it is possible to infer the range of conformations present in solution and hence the extent of disorder; n-ESI is highly sensitive to the degree of folding at the moment of transfer from the liquid to the gas phase. The combination of mass spectrometry with ion mobility (IM-MS) provides rotationally averaged collision cross-sections of molecular ions which can be correlated with conformation; this too can be applied to IDPs. Another feature which can be monitored by IM-MS is the tendency of disordered proteins to form amyloid fibrils, the protein aggregates involved in the onset of neurodegenerative diseases such as Parkinson's and Alzheimer's. IM-MS provides a useful insight into events that occur during the early stages of aggregation including delineating the structure of the monomer, identifying oligomer distributions, and revealing mechanistic details of the aggregation process. Here we will review the use of MS and IM-MS to study IDPs using examples from our own and other laboratories.

DOI: 10.1039/C2AN35665A

Effect of Protonation State on the Stability of Amyloid Oligomers Assembled from TTR(105-115)

M Porrini, U Zachariae, PE Barran & CE MacPhee

J. Phys. Chem. Lett. 4: 1233-1238 (2013)

Amyloid fibrils are self-assembled aggregates of polypeptides that are implicated in the development of several human diseases. A peptide derived from amino acids 105-115 of the human plasma protein transthyretin forms homogeneous and well-defined fibrils and, as a model system, has been the focus of a number of studies investigating the formation and structure of this class of aggregates. Self-assembly of TTR(105-115) occurs at low pH, and this work explores the effect of protonation on the growth and stability of small cross-β aggregates. Using molecular dynamics simulations of structures up to the decamer in both protonated and deprotonated states, we find that, whereas hexamers are more stable for protonated peptides, higher order oligomers are more stable when the peptides are deprotonated. Our findings imply a change in the acid pK of the protonated C-terminal group during the formation of fibrils, which leads to stabilization of higher-order oligomers through electrostatic interactions.

DOI: 10.1021/jz400372u

Gender differences in conceptual understanding of Newtonian mechanics: a UK cross-institution comparison

S Bates, R Donnelly, CE MacPhee, D Sands, M Birch and N R Walet

Eur. J. Phys. 34: 421-434 (2013)

We present the results of a combined study from three UK universities where we investigate the existence and persistence of a performance gender gap in conceptual understanding of Newtonian mechanics. Using the Force Concept Inventory, we find that students at all three universities exhibit a statistically significant gender gap, with males outperforming females. This gap is narrowed but not eliminated after instruction, using a variety of instructional approaches. Furthermore, we find that before instruction the quartile with the lowest performance on the diagnostic instrument comprises a disproportionately high fraction (~50%) of the total female cohort. The majority of these students remain in the lowest-performing quartile post-instruction. Analysis of responses to individual items shows that male students outperform female students on practically all items on the instrument. Comparing the performance of the same group of students on end-of-course examinations, we find no statistically significant gender gaps.

doi:10.1088/0143-0807/34/2/421

Atomic structure and hierarchical assembly of a cross-β amyloid fibril

Fitzpatrick AWP, Debelouchina GT, Bayro MJ, Clare DK, Caporini MA, Bajaj VS, Jaroniec CP, Wang L, Ladizhansky V, Müller SA, MacPhee CE, Waudby CA, Mott HR, De Simone A, Knowles TPJ, Saibil HR, Vendruscolo M, Orlova EV, Griffin RG, Dobson CM

Proc Nat'l Acad Sci USA 110: 5468-5473 (2013)

The cross-β amyloid form of peptides and proteins represents an archetypal and widely accessible structure consisting of ordered arrays of β-sheet filaments. These complex aggregates have remarkable chemical and physical properties, and the conversion of normally soluble functional forms of proteins into amyloid structures is linked to many debilitating human diseases, including several common forms of age-related dementia. Despite their importance, however, cross-β amyloid fibrils have proved to be recalcitrant to detailed structural analysis. By combining structural constraints from a series of experimental techniques spanning five orders of magnitude in length scale—including magic angle spinning nuclear magnetic resonance spectroscopy, X-ray fiber diffraction, cryoelectron microscopy, scanning transmission electron microscopy, and atomic force microscopy—we report the atomic-resolution (0.5 Å) structures of three amyloid polymorphs formed by an 11-residue peptide. These structures reveal the details of the packing interactions by which the constituent β-strands are assembled hierarchically into protofilaments, filaments, and mature fibrils.

doi: 10.1073/pnas.1219476110