Ageing and vision: structure, stability and function of lens crystallins.
ABSTRACT 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. Understanding how the specific interactions that confer intrinsic stability of the protein fold are combined with the stabilizing effect of protein assembly, and how the non-specific interactions and associations of the assemblies enable the generation of highly concentrated solutions, is thus of importance to understand the loss of transparency of the lens with age. Post-translational modification can have a major effect on protein stability but an emerging theme of the few studies of the effect of post-translational modification of the crystallins is one of solubility and assembly. Here we review the structure, assembly, interactions, stability and post-translational modifications of the crystallins, not only in isolation but also as part of a multi-component system. The available data are discussed in the context of the establishment, the maintenance and finally, with age, the loss of transparency of the lens. Understanding the structural basis of protein stability and interactions in the healthy eye lens is the route to solve the enormous medical and economical problem of cataract.
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ABSTRACT: Ultraviolet radiation is a risk factor for cataractogenesis. It is believed that enhanced rates of lens opacification and cataract formation are the results of gradual loss of chaperone-like efficiency of α-crystallin upon exposure to UV light. To characterize chaperone-like activity of α-crystallin damaged by UV irradiation, a test system based on dithiothreitol-induced aggregation of holo-α-lactalbumin from bovine milk was used. The adsorption capacity of α-crystallin (AC0) with respect to the target protein (α-lactalbumin) was used as a measure of anti-aggregation activity of α-crystallin. The data on SDS-PAGE testify that UV irradiation of α-crystallin results in covalent cross-linking of subunits in α-crystallin oligomers. The dependence of AC0 value on the irradiation dose was compared with the UV-induced diminution of the portion of native α-crystallin estimated from the data on differential scanning calorimetry. On the basis of such comparison a conclusion has been made that the loss in chaperone-like activity is mainly due to UV-induced denaturation of α-crystallin subunits. Cross-linking of remaining native subunits leads to an additional decrease in anti-aggregation activity.International Journal of Biological Macromolecules 11/2014; · 3.10 Impact Factor
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ABSTRACT: Introduction Cataract formation refers to the opacification of the eye lens and is a serious medical condition. Over 50% of humans over the age of 65 will develop cataracts which can result in total blindness. The mechanisms for cataract formation are poorly understood . Changes in the aggregation state of the eye lens proteins are responsible for cataract formation. One form of cataract formation occurs when the eye proteins reversibly aggregate upon cooling and the lens undergoes a reversible opacification . This effect has been studied in depth in mammals. While mammalian eyes exhibit cold cataract beginning at temperatures below 19ºC, the lens of the Antarctic fish, which lives at -2ºC are completely clear . In this study, we characterize differences in eye lens proteins to understand the variations in protein aggregation. Fish lenses are dense in protein to generate a refractive index large enough to focus light . The mechanism for focusing lies in changing the eye shape. For the bovine lens, protein densities are not as high, and focusing results from changes in lens shape. The dense protein state of the fish lens will result in large osmotic pressures, unless the proteins are attractive. The evolutionary drive to focus light in an aqueous environment has resulted in a balance of protein concentration and strength of attraction where the osmotic pressure of the cell matches the external osmotic pressure and lens transparency has been achieved. The bovine lens has been treated as a model system for physical and biochemical properties of the crystallin proteins, which are distinguished by size into: α, β and γ proteins. These proteins interact in complex manners such that the α and β proteins exist as aggregates whereas the γ proteins are monomeric. The γ proteins have been implicated in cold cataract formation. Even though there is substantially more γ in the fish, there is no evidence of cold cataract formation.
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ABSTRACT: Hyperthermostable proteins are highly resistant to various extreme conditions. Many factors have been proposed to contribute to their ultrahigh structural stability. Some thermostable proteins have larger oligomeric size when compared to their mesophilic homologues. The formation of compact oligomers can minimize the solvent accessible surface area and increase the changes of Gibbs free energy for unfolding. Similar to mesophilic proteins, hyperthermostable proteins also face the problem of unproductive aggregation. In this research, we investigated the role of high-order oligomerization in the fight against aggregation by a hyperthermostable superoxide dismutase identified from Tengchong, China (tcSOD). Besides the predominant tetramers, tcSOD could also form active high-order oligomers containing at least eight subunits. The dynamic equilibrium between tetramers and high-order oligomers was not significantly affected by pH, salt concentration or moderate temperature. The secondary and tertiary structures of tcSOD remained unchanged during heating, while cross-linking experiments showed that there were conformational changes or structural fluctuations at high temperatures. Mutational analysis indicated that the last helix at the C-terminus was involved in the formation of high-order oligomers, probably via domain swapping. Based on these results, we proposed that the reversible conversion between the active tetramers and high-order oligomers might provide a buffering system for tcSOD to fight against the irreversible protein aggregation pathway. The formation of active high-order oligomers not only increases the energy barrier between the native state and unfolded/aggregated state, but also provides the enzyme the ability to reproduce the predominant oligomers from the active high-order oligomers.PLoS ONE 10/2014; 9(10):e109657. · 3.53 Impact Factor