Flexible nanoassembly for sequestering non-native proteins

A crystal structure of a yeast small heat shock protein reported by Hanazono and colleagues in this issue of Structure reveals the versatility of the α-crystallin domain dimer for building assemblies of different size and symmetry. The domains assemble into a vessel filled with hydrophobic sequence extensions enriched with phenylalanines

Gamma-D crystallin gene (CRYGD) mutation causes autosomal dominant congenital cerulean cataracts

Congenital cataracts are a major cause of bilateral visual impairment in childhood. We mapped the gene responsible for autosomal congenital cerulean cataracts to chromosome 2q33-35 in a four generation family of Moroccan descent. The maximum lod score (7.19 at recombination fraction theta=0) was obtained for marker D2S2208 near the g-crystallin gene (CRYG) cluster. Sequencing of the coding regions of the CRYGA, B, C, and D genes showed the presence of a heterozygous C>A transversion in exon 2 of CRYGD that is associated with cataracts in this family. This mutation resulted in a proline to threonine substitution at amino acid 23 of the protein in the first of the four Greek key motifs that characterise this protein. We show that although the x ray crystallography modelling does not indicate any change of the backbone conformation, the mutation affects a region of the Greek key motif that is important for determining the topology of this protein fold. Our data suggest strongly that the proline to threonine substitution may alter the protein folding or decrease the thermodynamic stability or solubility of the protein. Furthermore, this is the first report of a mutation in this gene resulting in autosomal dominant congenital cerulean cataracts

Tracking socioeconomic vulnerability using network analysis: insights from an avian influenza outbreak in an ostrich production network

BACKGROUND: The focus of management in many complex systems is shifting towards facilitation, adaptation, building resilience, and reducing vulnerability. Resilience management requires the development and application of general heuristics and methods for tracking changes in both resilience and vulnerability. We explored the emergence of vulnerability in the South African domestic ostrich industry, an animal production system which typically involves 3-4 movements of each bird during its lifetime. This system has experienced several disease outbreaks, and the aim of this study was to investigate whether these movements have contributed to the vulnerability of this system to large disease outbreaks. METHODOLOGY/PRINCIPAL FINDINGS: The ostrich production system requires numerous movements of birds between different farm types associated with growth (i.e. Hatchery to juvenile rearing farm to adult rearing farm). We used 5 years of movement records between 2005 and 2011 prior to an outbreak of Highly Pathogenic Avian Influenza (H5N2). These data were analyzed using a network analysis in which the farms were represented as nodes and the movements of birds as links. We tested the hypothesis that increasing economic efficiency in the domestic ostrich industry in South Africa made the system more vulnerable to outbreak of Highly Pathogenic Avian Influenza (H5N2). Our results indicated that as time progressed, the network became increasingly vulnerable to pathogen outbreaks. The farms that became infected during the outbreak displayed network qualities, such as significantly higher connectivity and centrality, which predisposed them to be more vulnerable to disease outbreak. Conclusions/Significance Taken in the context of previous research, our results provide strong support for the application of network analysis to track vulnerability, while also providing useful practical implications for system monitoring and management

Wrapping the alpha-crystallin domain fold in a chaperone assembly

Small heat shock proteins (sHsps) are oligomers that perform a protective function by binding denatured proteins. Although ubiquitous, they are of variable sequence except for a C-terminal similar to 90-residue "alpha-crystallin domain". Unlike larger stress response chaperones, sHsps are ATP-independent and generally form polydisperse assemblies. One proposed mechanism of action involves these assemblies breaking into smaller subunits in response to stress, before binding unfolding substrate and reforming into larger complexes. Two previously solved non-metazoan sHsp multimers are built from dimers formed by domain swapping between the alpha-crystallin domains,. adding to evidence that the smaller subunits are dimers. Here, the 2.5 angstrom resolution structure of an sHsp from the parasitic flatworm Taenia saginata Tsp36, the first metazoan crystal structure, shows a new mode of dimerization involving N-terminal regions, which differs from that seen for non-metazoan sHsps. Sequence differences in the a-crystallin domains between metazoans and nonmetazoans are critical to the different mechanism of dimerization, suggesting that some structural features seen for Tsp36 may be generalized to other metazoan sHsps. The structure also indicates scope for flexible assembly of subunits, supporting the proposed process of oligomer breakdown, substrate binding and reassembly as the chaperone mechanism. It further shows how sHsps can bind coil and secondary structural elements by wrapping them around the alpha-crystallin domain. The structure also illustrates possible roles for conserved residues associated with disease, and suggests a mechanism for the sHsp-related pathogenicity of some flatworm infections. Tsp36, like other flatworm sHsps, possesses two divergent sHsp repeats per monomer. Together with the two previously solved structures, a total of four alpha-crystallin domain structures are now available, giving a better definition of domain boundaries for sHsps

Terminal regions confer plasticity to the tetrameric assembly of human HspB2 and HspB3

Heterogeneity in small heat shock proteins (sHsps) spans multiple spatiotemporal regimes – from fast fluctuations of part of the protein, to conformational variability of tertiary structure, plasticity of the interfaces, and polydispersity of the inter-converting, and co-assembling oligomers. This heterogeneity and dynamic nature of sHsps has significantly hindered their structural characterisation. Atomic-coordinates are particularly lacking for vertebrate sHsps, where most available structures are of extensively truncated homomers. sHsps play important roles in maintaining protein levels in the cell and therefore in organismal health and disease. HspB2 and HspB3 are vertebrate sHsps that are found co-assembled in neuromuscular cells, and variants thereof are associated with disease. Here, we present the structure of human HspB2/B3, which crystallised as a hetero-tetramer in a 3:1 ratio. In the HspB2/B3 tetramer, the four a-crystallin domains (ACDs) assemble into a flattened tetrahedron which is pierced by two non-intersecting approximate dyads. Assembly is mediated by flexible “nuts and bolts” involving IXI/V motifs from terminal regions filling ACD pockets. Parts of the N-terminal region bind in an unfolded conformation into the anti-parallel shared ACD dimer grooves. Tracts of the terminal regions are not resolved, most likely due to their disorder in the crystal lattice. This first structure of a full-length human sHsp heteromer reveals the heterogeneous interactions of the terminal regions and suggests a plasticity that is important for the cytoprotective functions of sHsps

Structure and evolution of crystallins

Book synopsis: As the first comprehensive reference for the eye, its support structures, diseases, and treatments, Encyclopedia of the Eye is an important resource for all visual scientists, ophthalmologists, and optometrists, as well as researchers in immunology, infectious disease, cell biology, neurobiology and related disciplines. This four-volume reference is unique in its coverage of information on all tissues important for vision, including the retina, cornea and lens. It also covers the physiological and pathophysiologic processes that affect all eye tissues. This Encyclopedia is invaluable for graduate students and postdoctoral fellows who are seeking an introduction to an area of eye research. Each chapter explains the basic concepts and provides references to relevant chapters within the Encyclopedia and more detailed articles across the wider research literature. The Encyclopedia is also particularly useful for visual scientists and practitioners who are researching a new area, seeking deeper understanding of important research articles in fields adjacent to their own, or reviewing a grant outside their immediate area of expertise

The Family of Small Heat Shock Proteins: Assembly and Binding Functions

Small heat shock proteins (sHsps) are a diverse family evolved from ancient stress proteins that are now associated with protecting microbial, plant, worm, insect and mammalian cells under stress, where prevention of aggregation of destabilized proteins is considered to be their general function. The sequences all contain a short a-crystallin domain (ACD) bracketed by variable sequence extensions that contribute to a dynamic continuum of oligomeric forms. A few large symmetric oligomeric assemblies from nonmetazoans have been resolved by crystallography, as well as a few metazoan subassembly forms. A structure-based sequence alignment of the ACDs from resolved structures with representative sequences of sHsps from species of high economic, model or medical value is presented. The ACDs are portrayed to emphasize their interaction interfaces. The construction of a range of nanoassemblies from a polyvalent building block by binding IXI/V sequence motifs in extensions into partner ACD pockets is illustrated. This interaction mechanism can also be used to connect sHsps to energy driven chaperone machines, as shown for the interaction with cochaperone Bag3 in myofibril Z-disks. The resolved complete assemblies are built from a strand exchange dimer, whereas animal sHsps have an antiparallel (AP) dimer interface that forms a shared groove which can change shape. Spectroscopy has shown how the AP interface is regulated by pH. Animal sHsp full assemblies are undetermined, although some clues as to the structural role of the hydrophobic N-terminal extensions can be surmised from analysis of the resolved assemblies. A mechanism for prevention of protein aggregation is likely to involve a stress-regulated ensemble of interconverting destabilized sHsp assemblies when interfaces and extensions become more accessible to the proteome. Rapid developments in EM offer the possibility of resolution of an sHsp ensemble if the level of polydispersity can be lowered

sHSP in the eye lens: crystallin mutations, cataract and proteostasis

Alpha-crystallin, a major component of the eye lens cytoplasm, is a large multimer formed from two members of the small heat shock protein (sHsp) family. Inherited crystallin mutations are a common cause of childhood cataract, whereas miscellaneous changes to the long-lived crystallins cause age-related cataract, the most common cause of blindness worldwide. Newly formed eye lens cells use proteostasis to deal with the consequences of mutations, whereas mature lens cells, devoid of the ATP-driven folding and degradation machines, are hypothesised to have the α-crystallin “holdase” chaperone function to prevent protein aggregation. We discuss the impact of truncating and missense mutations on α-crystallin, based on recent progress towards determining sHsp 3D structure. Dominant missense mutations to the “α-crystallin domain” of αA- (HSPB4) or αB-crystallin (HSPB5) occur on residues predicted to facilitate domain dynamics. αB-crystallin is also expressed in striated muscle and mutations cause myopathy. The impact on these cellular cytoplasms is compared where sHsp multimer partners and metabolic constraints are different. Selected inherited mutations of the lens β- and γ-crystallins are considered in the context of their possible dependence on the “holdase” chaperone function of α-crystallin. Looking at discrete changes to specific crystallin polypeptide chains that can function as chaperone or substrate provide insights into the workings of a cytoplasmic proteostatic system. These observations provide a framework for validating the function of α-crystallin as a chaperone, or as a lens space filler adapted from a chaperone function. Understanding the mechanistic role of α-crystallins will aid progress in research into age-related cataract and adult-onset myopathy