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 characterization. Atomic coordinates are particularly lacking for vertebrate sHsps, where most available structures are of extensively truncated homomers.

Functions of crystallins in and out of lens: roles in elongated and post-mitotic cells.

The vertebrate lens evolved to collect light and focus it onto the retina. In development, the lens grows through massive elongation of epithelial cells possibly recapitulating the evolutionary origins of the lens. The refractive index of the lens is largely dependent on high concentrations of soluble proteins called crystallins.

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.

Evolution of crystallins for a role in the vertebrate eye lens.

The camera eye lens of vertebrates is a classic example of the re-engineering of existing protein components to fashion a new device. The bulk of the lens is formed from proteins belonging to two superfamilies, the α-crystallins and the βγ-crystallins. Tracing their ancestry may throw light on the origin of the optics of the lens.

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

α-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 hypothesized to have the α-crystallin "holdase" chaperone function to prevent protein aggregation.

Crystal structure of R120G disease mutant of human αB-crystallin domain dimer shows closure of a groove.

Small heat shock proteins form large cytosolic assemblies from an "α-crystallin domain" (ACD) flanked by sequence extensions. Mutation of a conserved arginine in the ACD of several human small heat shock protein family members causes many common inherited diseases of the lens and neuromuscular system. The mutation R120G in αB-crystallin causes myopathy, cardiomyopathy and cataract.

Crystal structures of alpha-crystallin domain dimers of alphaB-crystallin and Hsp20.

Small heat shock proteins (sHsps) are a family of large and dynamic oligomers highly expressed in long-lived cells of muscle, lens and brain. Several family members are upregulated during stress, and some are strongly cytoprotective. Their polydispersity has hindered high-resolution structure analyses, particularly for vertebrate sHsps.

Association of partially folded lens betaB2-crystallins with the alpha-crystallin molecular chaperone.

Age-related cataract is a result of crystallins, the predominant lens proteins, forming light-scattering aggregates. In the low protein turnover environment of the eye lens, the crystallins are susceptible to modifications that can reduce stability, increasing the probability of unfolding and aggregation events occurring. It is hypothesized that the alpha-crystallin molecular chaperone system recognizes and binds these proteins before they can form the light-scattering centres that result in cataract, thus maintaining the long-term transparency of the lens.

Biophysical properties of gammaC-crystallin in human and mouse eye lens: the role of molecular dipoles.

The eye lens is packed with soluble crystallin proteins, providing a lifetime of transparency and light refraction. gamma-Crystallins are major components of the dense, high refractive index central regions of the lens and generally have high solubility, high stability and high levels of cysteine residues. Human gammaC belongs to a group of gamma-crystallins with a pair of cysteine residues at positions 78 and 79.

A reference dataset for circular dichroism spectroscopy tailored for the betagamma-crystallin lens proteins.

Circular dichroism (CD) spectroscopy is a powerful solution technique for the study of protein secondary structure. As hierarchical euclidean clustering analyses of high quality crystallin synchrotron radiation circular dichroism (SRCD) spectral data can be separated into structural groups based solely on spectral information, the technique can potentially be improved to more accurately determine secondary structures and monitor conformational changes in crystallins. Secondary structure estimates can be determined through use of reference datasets of circular dichroism spectra from proteins with determined crystal structures.

Mutation of interfaces in domain-swapped human betaB2-crystallin.

The superfamily of eye lens betagamma-crystallins is highly modularized, with Greek key motifs being used to form symmetric domains. Sequences of monomeric gamma-crystallins and oligomeric beta-crystallins fold into two domains that pair about a further conserved symmetric interface. Conservation of this assembly interface by domain swapping is the device adopted by family member betaB2-crystallin to form a solution dimer.

Lengsin is a survivor of an ancient family of class I glutamine synthetases re-engineered by evolution for a role in the vertebrate lens.

Lengsin is a major protein of the vertebrate eye lens. It belongs to the hitherto purely prokaryotic GS I branch of the glutamine synthetase (GS) superfamily, but has no enzyme activity. Like the taxon-specific crystallins, Lengsin is the result of the recruitment of an ancient enzyme to a noncatalytic role in the vertebrate lens.

In vivo heteromer formation. Expression of soluble betaA4-crystallin requires coexpression of a heteromeric partner.

The beta-crystallins are a family of long-lived, abundant structural proteins that are coexpressed in the vertebrate lens. As beta-crystallins form heteromers, a process that involves transient exposure of hydrophobic interfaces, we have examined whether in vivobeta-crystallin assembly is enhanced by protein chaperones, either small heat shock proteins, Hsp27 or alphaB-crystallin, or Hsp70. We show here that betaA4-crystallin is abundantly expressed in HeLa cells, but rapidly degraded, irrespective of the presence of Hsp27, alphaB-crystallin or Hsp70.

Urochordate betagamma-crystallin and the evolutionary origin of the vertebrate eye lens.

A refracting lens is a key component of our image-forming camera eye; however, its evolutionary origin is unknown because precursor structures appear absent in nonvertebrates. The vertebrate betagamma-crystallin genes encode abundant structural proteins critical for the function of the lens. We show that the urochordate Ciona intestinalis, which split from the vertebrate lineage before the evolution of the lens, has a single gene coding for a single domain monomeric betagamma-crystallin.

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 approximately 90-residue "alpha-crystallin domain". Unlike larger stress response chaperones, sHsps are ATP-independent and generally form polydisperse assemblies.

gammaN-crystallin and the evolution of the betagamma-crystallin superfamily in vertebrates.

The beta and gamma crystallins are evolutionarily related families of proteins that make up a large part of the refractive structure of the vertebrate eye lens. Each family has a distinctive gene structure that reflects a history of successive gene duplications. A survey of gamma-crystallins expressed in mammal, reptile, bird and fish species (particularly in the zebrafish, Danio rerio) has led to the discovery of gammaN-crystallin, an evolutionary bridge between the beta and gamma families.

Unfolding crystallins: the destabilizing role of a beta-hairpin cysteine in betaB2-crystallin by simulation and experiment.

The thermodynamic and kinetic stabilities of the eye lens family of betagamma-crystallins are important factors in the etiology of senile cataract. They control the chance of proteins unfolding, which can lead to aggregation and loss of transparency. betaB2-Crystallin orthologs are of low stability and comprise two typical betagamma-crystallin domains, although, uniquely, the N-terminal domain has a cysteine in one of the conserved folded beta-hairpins.

Specific interaction between lens MIP/Aquaporin-0 and two members of the gamma-crystallin family.

Purpose: Major Intrinsic Protein (MIP)/Aquaporin 0 is required for lens transparency and is specifically expressed in lens fiber cell membranes. We have demonstrated previously that in the rat lens MIP interacts specifically with gammaE-crystallin, resulting in its recruitment to the plasma membrane. Our goal was to examine the interaction or lack of interaction between MIP and all members of the gamma-crystallin family and to provide evidence for a physiological role these interactions may play in gamma-crystallin or MIP function.

Sulfur in human crystallins.

Molecular models of human gamma-crystallins and the 'alpha-crystallin domain' of human alphaA-crystallin have been built based on available related X-ray crystal structures. The accessibilities of the component cysteine, methionine and tryptophan side chains in the crystallin models have been calculated. The reactivities of these cysteines, which are oxidised in cataract, are assessed based on their known modifications and within the context of their location within the 3D models.

The P23T cataract mutation causes loss of solubility of folded gammaD-crystallin.

Mutations in the human gammaD-crystallin gene have been linked to several types of congenital cataracts. In particular, the Pro23 to Thr (P23T) mutation of human gammaD crystallin has been linked to cerulean, lamellar, coralliform, and fasciculiform congenital cataracts. We have expressed and purified wild-type human gammaD, P23T, and the Pro23 to Ser23 (P23S) mutant.

Ageing and vision: structure, stability and function of lens crystallins.

The alpha-, beta- and gamma-crystallins are the major protein components of the vertebrate eye lens, alpha-crystallin as a molecular chaperone as well as a structural protein, beta- and gamma-crystallins as structural proteins. For the lens to be able to retain life-long transparency in the absence of protein turnover, the crystallins must meet not only the requirement of solubility associated with high cellular concentration but that of longevity as well. For proteins, longevity is commonly assumed to be correlated with long-term retention of native structure, which in turn can be due to inherent thermodynamic stability, efficient capture and refolding of non-native protein by chaperones, or a combination of both.

Characterization of the G91del CRYBA1/3-crystallin protein: a cause of human inherited cataract.

Congenital cataract is a leading cause of visual disability in children. Inherited isolated (non-syndromic) cataract represents a significant proportion of cases and the identification of genes responsible for inherited cataract will lead to a better understanding of the mechanism of cataract formation at the molecular level both in congenital and age-related cataract. Crystallins are abundantly expressed in the developing human lens and represent excellent candidate genes for inherited cataract.