Glycosylation differentially modulates membranolytic and chaperone-like activities of PDC-109, the major protein of bovine seminal plasma.

The major bovine seminal plasma protein, PDC-109, is a mixture of glycosylated (BSP-A1) and non-glycosylated (BSP-A2) isoforms of a 109-residue long polypeptide. It binds to spermatozoa by specifically recognizing choline phospholipids on the plasma membrane and destabilizes it by penetrating the hydrophobic interior, resulting in lipid efflux, which is necessary for sperm capacitation and successful fertilization. PDC-109 also acts as a molecular chaperone and protects target proteins from denaturation and aggregation induced by various types of stress. In order to investigate the role of glycosylation in these activities, we have separated BSP-A1 and BSP-A2 from PDC-109, and also cloned and expressed BSP-A2 in E. coli and purified the recombinant BSP-A2 (rBSP-A2) to homogeneity. Employing biophysical and biochemical approaches we have investigated the membrane-perturbing and chaperone-like activities (CLA) of PDC-109, BSP-A1, BSP-A2 and recombinant BSP-A2 (rBSP-A2). The results obtained demonstrate that glycan-lacking wild-type BSP-A2 and rBSP-A2 exhibit higher membrane-perturbing activity but decreased CLA as compared to PDC-109. In contrast, BSP-A1 exhibits significantly higher CLA than PDC-109, but its ability to destabilize membranes is considerably lower. This differential modulation of the membrane-perturbing and chaperone-like activities has been explained on the basis of higher membrane-penetrating ability and lower solubility of glycan-lacking BSP-A2 as compared to the glycosylated BSP-A1.

The Local Effect of Alendronate with Intra-alveolar Collagen Sponges on Post Extraction Alveolar ridge Resorption: A Clinical Trial.

INTRODUCTION: Extraction of teeth is followed by resorption of the residual alveolar ridge that continues throughout life resulting in loss of alveolar height and width. Of the numerous techniques that have been used to arrest post extraction alveoloar ridge resorption, the placement of a graft material inside the socket immediately after extraction has been mostly followed. Type 1 collagen is one of the commonly used graft material that prevent resorption by providing dimensional stability to the socket. Bisphosphonates are an anti-osteoclastic drug that prevent resorption by disrupting the membrane ruffling of the osteoclasts. Alendronate a bisphosphonate, is primarily used in diseases with bone loss. It has been used to reduce active bone resorption significantly without interfering with bone mineralization and quality. The need for the study is to examine the inhibitory effect of alendronate on residual ridge resorption when applied locally in combination with type I collagen on alveolar bone immediately following tooth extraction. MATERIALS AND METHODS: Twenty patients with age between 30 and 65 years were selected from the out patient department of The Oxford Dental College and Hospital. The patients were divided into two groups. In the first group after extraction of teeth from premolar to midline the sockets were irrigated with saline and sutured. On the left side type I collagen sponge was placed and sutured. In the other group the right side was treated the same way after extraction as in first group where as in the left side sockets type I collagen soaked in 20 mg/ml of alendronate was placed and sutured. Patients were evaluated clinically for any local irritation as well as radiologically with orthopantomograph X-rays were taken immediately after the extraction, 1 month after extraction and 4 months after extraction to determine the amount of bone loss prevented. RESULTS: The statistically significant bone loss prevented by the collagen alone was 22.8 % and in collagen with alendronate group was 44.38 % at the end of 4 months. CONCLUSION: Type I collagen soaked with alendronate when placed in the socket immediately after extraction of teeth prevents post-extraction alveolar ridge resorption.

Effects of tetracycline-containing gel and a mixture of tetracycline and citric acid-containing gel on non-surgical periodontal therapy.

AIMS AND OBJECTIVES: The purpose of this study was to assess the clinical and microbiological effects of a newly developed root-conditioning gel system containing tetracycline and a mixture of tetracycline and citric acid on non-surgical periodontal therapy. MATERIALS AND METHODS: Four anterior teeth from four quadrants with a probing depth of 4-6 mm, in each of the 20 subjects with chronic periodontitis, were subjected to four different modalities of treatment. A total of 80 teeths were divided into four groups of 20 teeth each taken from separate quadrants, on the basis of one of the following four treatments: (1) Root planning alone in first quadrant (RP group); (2) tetracycline-containing gel in the second quadrant (TCG group); (3) root planning plus tetracycline-containing gel in third quadrant (RP + TCG group); (4) root planning plus a mixture of tetracycline and citric acid-containing gel in fourth quadrant (RP + TC-CAG group). Plaque index (PI), sulcular bleeding index, probing pocket depth, and clinical attachment level were measured for 0 day, 8 th week, and 12 th week, respectively. Subgingival plaque samples from each site were collected at the same visits and examined with dark field microscope for proportions of motile rods and spirochetes. RESULTS: From 0 day to 12 th week, PI, sulcular bleeding index, probing pocket depth, and clinical attachment levels decreased significantly in all the groups. From 0 day to 12 th week, RP + TC-CAG group showed a significantly higher change in the PI score. From 0 day to 12 th week, RP group showed a significantly higher change in sulcular bleeding index score. A significant decrease in probing pocket depth and gain in clinical attachment level was noted at 12 th week in RP + TC-CAG group compared to the other groups. A significant decrease in the proportion of motile rods was found primarily in the RP + TC-CAG group. There was a decrease in the proportion of spirochetes in all the groups. CONCLUSION: The results indicated that the use of a mixture of tetracycline and citric acid-containing gel was effective in improving gingival health and in changing subgingival microflora.

Bone regeneration with plasma-rich-protein following enucleation of traumatic bone cyst.

Traumatic bone cyst is an uncommon non-epithelium lined cavity and is seen frequently in young individuals. The lesion occurs more commonly in the mandible, involving the posterior region. It is generally asymptomatic and is diagnosed on routine radiographic examination. The cystic cavity is usually empty and there is scanty material for histological examination. Surgical curettage is usually done and recurrence is rare. A case of traumatic bone cyst occurring in the anterior region of mandible in a young boy is presented. Following surgical intervention, plasma-rich-protein was placed in the cystic cavity. The lesion showed progressive resolution and bone regeneration of the cystic cavity within a short period of time.

Effectiveness of a controlled release chlorhexidine chip (PerioCol™-CG) as an adjunctive to scaling and root planing when compared to scaling and root planing alone in the treatment of chronic periodontitis: A comparative study.

AIMS AND OBJECTIVES: The aim of this study is to evaluate the effectiveness of a controlled-release chlorhexidine chip as an adjunctive therapy to scaling and root planing when compared with scaling and root planing alone in the treatment of chronic periodontitis. MATERIALS AND METHODS: 20 patients with a total number of 40 posterior sites were selected. These sites were divided into two groups in a split mouth design,: Group A (control site) had 20 sites treated with scaling and root planing alone and Group B (test site) had 20 sites treated with scaling and root planing and PerioCol™-CG. The clinical parameters (Plaque index, bleeding on probing, probing pocket depth, clinical attachment level) were recorded at baseline, 90(th) and 180(th) day for both the groups. RESULTS: When both groups were compared the change in Plaque index was significantly higher in Group B when compared to Group A on the 90(th) day and 180(th) day. However, there was no statistically significant difference in the mean percentage of gingival bleeding sites between the two groups on the 90(th) day, though Group B showed a statistically higher reduction in the mean percentage of gingival bleeding sites at the end of 180(th) day. There was no statistically significant difference in probing pocket depth between the two groups on both 90(th) and 180(th) day. Gain in clinical attachment level was significantly higher in Group B when compared to Group A on the 90(th) and 180(th) day. CONCLUSION: From the results observed in this study, it can be concluded that the adjunctive use of PerioCol™-CG was safe and provided significant improvement in both Plaque index and gingival bleeding index. It was also more favorable than scaling and root planing alone for gain in clinical attachment level.

Utility of the Malayalam translation of the 7- minute screen for Alzheimer's disease risk in an Indian community.

BACKGROUND: Alzheimer's disease (AD) is suspected to be currently under-diagnosed in India, thus the need for a brief, effective screening test for the condition. AIMS: We aimed to test the Malayalam translation of the 7-Minute Screen (7MS) for detecting those at high risk for AD and to report on the subscores used to derive the Alzheimer's risk score. SETTING AND DESIGN: This study was performed in Kerala State amongst young university students and elders in residential care homes. MATERIALS AND METHODS: Two hundred and eighty-two volunteers were tested, 178 young controls (aged 20-29) and 104 literate elders, (55-92 years). None were clinically diagnosed with AD. STATISTICAL ANALYSES: Elders and controls were assessed as High or Low AD Risk with the published 7MS algorithm. Performance was compared between groups with ANOVA. RESULTS: The algorithm estimated high (n=61/104) or low (n=40/104) AD risk in the elderly. Significant differences were found between controls, low- and high-risk groups on all four components of the screen (Orientation: F=131.1, Enhanced Cued Recall: F=23.4, Clock Drawing: F=65.1, Verbal Fluency: F=15.7, P<0.0001 for all) and in the risk scores (F=144.7, P<0.0001). Age and gender affected verbal fluency, orientation and clock drawing performance. The high-risk group had worse scores for orientation and better scores for memory than previously reported for AD cases in other populations. CONCLUSIONS: The 7MS may be a useful screening test for cognitive impairment in India. Suggestions are given for revising the 'risk algorithm' for more appropriate AD risk assessment in this population.

Autocatalytic alkyne cycloadditions: evidence for colloidal Pt catalysis.

Treatment of the four-membered platinacycle L2Pt(1,8-naphthalendiyl) (1) or the five-membered platinacycle L2Pt(1,12-triphenylendiyl) with excess PhCCPh at 120-150 degrees C gives the coupling products 1,2-diphenylacenaphthalene or 4,5-diphenylbenzo[e]pyrene and the alkyne complex L2Pt(eta2-PhCCPh). Both reactions show an accelerating rate, which has been traced to catalysis of the reaction by colloidal platinum formed by the reaction of O2 with L2Pt(eta2-PhCCPh).

Arginine hydrochloride enhances the dynamics of subunit assembly and the chaperone-like activity of alpha-crystallin.

PURPOSE: Alpha-crystallin, a major eye lens protein, bears homology with small heat shock proteins (sHsps) and exhibits molecular chaperone-like activity. Structural perturbation by temperature or low concentrations of denaturants leads to enhancement of its chaperone-like activity. We have earlier demonstrated similar enhancement of chaperone-like activity using biologically compatible solutes such as arginine hydrochloride and aminoguanidine. The purpose of the present study is to get an insight into the mechanism of the arginine induced enhancement of chaperone-like activity of alpha-crystallin. METHODS: The effect of arginine hydrochloride on the chaperone-like activity of alpha-crystallin at 25 degrees C was studied using DTT induced aggregation of insulin as a model system. Changes in the accessibility of the thiol group near the end of the alpha-crystallin domain in the absence and the presence of arginine hydrochloride were studied using dithiobisnitrobenzoic acid. Fluorescence resonance energy transfer studies were performed to investigate changes in the dynamics of the subunit assembly. Urea induced denaturation studies of alpha-crystallin were carried out to investigate structural destabilization of alpha-crystallin, if any, in the presence of arginine hydrochloride. RESULTS: Arginine hydrochloride increases the chaperone-like activity of alpha-crystallin several fold towards DTT induced aggregation of insulin at room temperature. Our study shows that both the extent and the rate of accessibility of the thiol group are increased in the presence of arginine. Fluorescence resonance energy transfer experiments show that arginine hydrochloride significantly increases the subunit exchange between the oligomers of alpha-crystallin. Arginine induced structural perturbation and loosening of subunit assembly of alpha-crystallin leads to overall destabilization of the protein as reflected by the urea denaturation study. CONCLUSIONS: Arginine perturbs the tertiary and quaternary structure of alpha-crystallin and enhances the dynamics of the subunit assembly leading to enhanced chaperone-like activity. Thus, in addition to size, surface hydrophobicity, and charge distribution, the dynamics of the subunit assembly appears to be one of the critical factors that can modulate the chaperone activity.

Effect of pyridoxine deficiency on the structural and functional development of hippocampus.

In this study it was attempted to understand the effect of pyridoxine deficiency on the structural and functional development of the hippocampus. Hippocampus has been closely associated with complex neuroendocrine control of physiological activities as well as behavioural responses including learning process and memory retention. Prenatal, preweanling and weanling deficiency of pyridoxine was induced in the experimental rats by feeding dams with diet deficient in pyridoxine during pregnancy and lactation. The general growth profile for pyridoxine deficient (PD) rats is compared with control ones. The structural changes in the hippocampus of pyridoxine deficient rats was investigated using the histological techniques. Hippocampal electrical activity was recorded from in vitro brain slice preparation. The study clearly showed the structural impairment in the hippocampus of PD rats. These anatomic anomalies might be related to poor neurointegrative development and neurophysiological deficits that occur in young one. The electrical activity recorded from hippocampal slices of PD rats showed significant variation when compared to controls. Pyridoxine deficiency is common in pregnant women who used anovulatory steroids before pregnancy. The pyridoxine deficiency of the mother may result in permanent behavioural abnormality and intellectual deficit in the progeny.

Naphthalenes, isoquinolines, and a benzazocine from zirconocene-copper-mediated coupling of benzocyclobutadiene with nitriles and alkynes.

[reaction: see text] Commercially available 1-bromobenzocyclobutene is a potentially useful synthon particularly with the application of organometallic methodology. Here we show that it is readily converted into Cp(2)Zr(benzocyclobutadiene), which couples with alkynes or nitriles giving five-membered zirconacycles. Treatment of these alkyne- or nitrile-derived zirconacycles with CuCl yields substituted naphthalenes, isoquinolines, or in the presence of MeO(2)C-CC-CO(2)Me, a 3-benzazocine containing an eight-membered ring [corrected].

Loss of dendritic connectivity in CA1, CA2, and CA3 neurons in hippocampus in rat under aluminum toxicity: antidotal effect of pyridoxine.

Aluminum chloride (AlCl(3); 4 mg/kg) was injected into the cerebrospinal fluid of adult rats as a one time dose. Rapid Golgi stained sections of hippocampus were examined for detailed histology of neurons in CA1, CA2, and CA3 areas. The axonal length and number of dendritic branches were seen reduced 30 days later in aluminum (Al)-injected group when compared to vehicle-injected controls. Of these perturbations, dendritic branches were seen reduced significantly. Al toxicity apparently affects neuronal connectivity in hippocampus. These perturbations are reversed by supplementing the feed with pyridoxine (8 mg/kg) for 30 days. As the loss of synaptic connectivity is a predominant feature of neurodegenerative disorders such as Alzheimer disease, this study may have implications in such disorders. Pyridoxine may be considered as a potent antidote to Al toxicity and neurodegenerative disorders such as Alzheimer disease.

Unfolding and refolding of a quinone oxidoreductase: alpha-crystallin, a molecular chaperone, assists its reactivation.

alpha-Crystallin, a member of the small heat-shock protein family and present in vertebrate eye lens, is known to prevent the aggregation of other proteins under conditions of stress. However, its role in the reactivation of enzymes from their non-native inactive states has not been clearly demonstrated. We have studied the effect of alpha-crystallin on the refolding of zeta-crystallin, a quinone oxidoreductase, from its different urea-denatured states. Co-refolding zeta-crystallin from its denatured state in 2.5 M urea with either calf eye lens alpha-crystallin or recombinant human alpha B-crystallin could significantly enhance its reactivation yield. alpha B-crystallin was found to be more efficient than alpha A-crystallin in chaperoning the refolding of zeta-crystallin. In order to understand the nature of the denatured state(s) of zeta-crystallin that can interact with alpha-crystallin, we have investigated the unfolding pathway of zeta-crystallin. We find that it unfolds through three distinct intermediates: an altered tetramer, a partially unfolded dimer, which is competent to fold back to its active state, and a partially unfolded monomer. The partially unfolded monomer is inactive, exhibits highly exposed hydrophobic surfaces and has significant secondary structural elements with little or no tertiary structure. This intermediate does not refold into the active state without assistance. alpha-Crystallin provides the required assistance and improves the reactivation yield several-fold.

Studies on the alpha-crystallin target protein binding sites: sequential binding with two target proteins.

PURPOSE: alpha-Crystallin belongs to a class of small heat shock proteins and is shown to prevent aggregation of several proteins. We have shown that the temperature-induced structural perturbation leads to several fold enhanced activity. The purpose of this study was to investigate the availability and specificity of the hydrophobic sites that might become available at elevated temperatures. Specifically, we address the following question: Is there an increased exposure of fixed number of hydrophobic sites as a function of temperature or does a new set of sites become available at elevated temperatures? METHODS: alpha-Crystallin target protein complexes were made at two different temperatures and this complex was investigated for its chaperone-like activity towards the same target protein and also other target proteins. DTT-induced aggregation of insulin, alpha-lactalbumin, thermal aggregation of betaL- and gamma-crystallin, and photo-aggregation of gamma-crystallin were used as model systems. Increased light scattering was used to monitor the progress of aggregation. RESULTS: alpha-Crystallin target protein complex prepared at 37 degrees C temperature was effective against thermal aggregation of betaL-crystallin as well as non-thermal aggregation at elevated temperatures. However, the complex prepared at high temperature was ineffective at lower temperatures as well as with other target proteins at both temperatures. CONCLUSIONS: More target protein binding sites become available at elevated temperatures. The sites available at low temperature are a subset of the total sites available at elevated temperatures.

Interaction of human recombinant alphaA- and alphaB-crystallins with early and late unfolding intermediates of citrate synthase on its thermal denaturation.

We have investigated the role of recombinant human alphaA- and alphaB-crystallins in the heat-induced inactivation and aggregation of citrate synthase. Homo-multimers of both alphaA- and alphaB-crystallins confer protection against heat-induced inactivation in a concentration-dependent manner and also prevent aggregation. Interaction of crystallins with early unfolding intermediates of citrate synthase reduces their partitioning into aggregation-prone intermediates. This appears to result in enhanced population of early unfolding intermediates that can be reactivated by its substrate, oxaloacetate. Both these homo-multimers do not form a stable complex with the early unfolding intermediates. However, they can form a soluble, stable complex with aggregation-prone late unfolding intermediates. This soluble complex formation prevents aggregation. Thus, it appears that the chaperone activity of alpha-crystallin involves both transient and stable interactions depending on the nature of intermediates on the unfolding pathway; one leads to reactivation of the enzyme activity while the other prevents aggregation.

Redox-regulated chaperone function and conformational changes of Escherichia coli Hsp33.

We have studied the chaperone activity and conformation of Escherichia coli heat shock protein (Hsp)33, whose activity is known to be switched on by oxidative conditions. While oxidized Hsp33 completely prevents the heat-induced aggregation of zeta-crystallin at 42 degrees C at a ratio of 1:1 (w/w), the reduced form exhibits only a marginal effect on the aggregation. Far UV-circular dichroism (CD) spectra show that reduced Hsp33 contains a significant alpha-helical component. Oxidation results in significant changes in the far UV-CD spectrum. Near UV-CD spectra show changes in tertiary structural packing upon oxidation. Polarity-sensitive fluorescent probes report enhanced hydrophobic surfaces in the oxidized Hsp33. Our studies show that the oxidative activation of the chaperone function of Hsp33 involves observable conformational changes accompanying increased exposure of hydrophobic pockets.

Enzymatic, clinical and histologic evaluation of corneal tissues in experimental fungal keratitis in rabbits.

Mycotic keratitis, being frequently refractive to most of the currently available antifungal therapy, continues to pose a therapeutic challenge to the clinician. In keratitis of infectious etiology stromal dissolution may be brought about by a combination of agent and host factors. An understanding of the source and nature of corneal tissue damage is essential for evolving more effective therapeutic modalities in the treatment of fungal keratitis. In the present study, we have characterized the extracellular proteases produced in vitro by corneal fungal pathogens namely the Aspergillus flavus and Fusarium solani when collagen was provided as the sole nitrogen source. In addition, fungal infected rabbit corneas were investigated for proteolytic activities and nature of inflammatory reaction. Gelatin zymography detected protease bands with molecular mass ranging from 100 to 200 kDa in the culture extracts of A. flavus, and a single major band of molecular mass approximately 200 kDa in the culture extracts of F. solani. A basal proteolytic activity of mass 65 kDa was visualized in all uninfected and infected rabbit corneal extracts. Infected corneas in addition revealed the presence of additional proteolytic species of mass 92 and 200 kDa. The enzyme inhibitory profile suggested that fungal cultures in vitro contained predominantly serine protease activity and to a lesser extent metalloprotease activity. However, fungal infected corneal homogenates showed the presence of metalloproteinase activity alone, the enzymatic activities entirely being sensitive to ethylene diamine tetra acetate (EDTA), a metalloprotease inhibitor. Interestingly, the serine proteolytic activity detected in fungal cultures in vitro was not present in the fungal infected corneas in vivo. However, the possible role of fungal serine proteases in the activation of corneal matrix metalloproteinases (MMPs) cannot be ruled out. Based on the criteria of molecular mass, proteolytic activity in the presence of calcium at neutral pH, and sensitivity to inhibition by a metalloprotease inhibitor, the 65 and 92 kDa gelatinases were identified as MMP 2 and MMP 9, respectively. The expression of 92 and 200 kDa gelatinases correlated positively with the amount of polymorphonuclear cells present in the infected tissues. Activated resident corneal cells or inflammatory cells may largely contribute to the increased proteolytic activities in fungal infected corneas resulting in tissue matrix degradation in fungal keratitis.

Vitamins and brain development.

Effects of deficiency of vitamins on early development of brain have been reviewed. Unusual developmental problems in neurogenesis specific for the brain and impairment of its functional capacities due to vitamin deficiency have been discussed. The species-specific "critical periods" in development of various systems have been mentioned. Indices such as reflex activity, locomotion, special senses, cognition and adaptive behavior were used for assessing brain maturation in experimental models and humans. Significant examples include brain anomalies in humans and other mammals caused by retinoid excess or deficit; increase in calbindin D28K, a vitamin D dependent calcium-binding protein during postnatal period in rat; hydrocephalus and exencephaly in prenatal rats and subarachnoidal or intracerebral hemorrhage in infants caused by vitamin E deficiency. Peripheral neuropathic lesions leading to infantile beriberi is caused by thiamine deficiency. Impaired growth in retinal layers leading to delay in maturation of electroretinogram and depth-perception in postnatal rats occur due to pyridoxine deficiency. Infants of severely vitamin B12 deficient mothers show abnormalities in behavior involving basal ganglia and pyramidal tract. Folic acid deficiency results in delayed maturation of the basic electroencepalographic patterns. In addition, vitamin-interactions leading to developmental errors have been pointed out. Vitamin B6 deficiency impairs vitamin B12 absorption and biotin deficiency may be aggravated by pantothenic acid deficiency. Vitamin C deficiency resulting in impaired metabolism may produce symptoms of deficiency of folic acid. Another characteristic examples is that iron absorption from dietary sources is dependent on ascorbic acid.

Detection and assay of proteases using calf lens beta-crystallin aggregate as substrate.

The eye lens protein, betaL-crystallin, aggregates and yields a turbid solution upon refolding from its denatured state. We have observed that the addition of trace amounts of protease results in clearing of this turbidity. Based on this observation, we have developed a simple and rapid method for the detection and assay of proteases. This assay can be performed in the pH range of 6.0-9.0. We could assay the activity of trypsin at a concentration as low as 5 microg/ml.

Structural and functional consequences of the mutation of a conserved arginine residue in alphaA and alphaB crystallins.

A point mutation of a highly conserved arginine residue in alphaA and alphaB crystallins was shown to cause autosomal dominant congenital cataract and desmin-related myopathy, respectively, in humans. To study the structural and functional consequences of this mutation, human alphaA and alphaB crystallin genes were cloned and the conserved arginine residue (Arg-116 in alphaA crystallin and Arg-120 in alphaB crystallin) mutated to Cys and Gly, respectively, by site-directed mutagenesis. The recombinant wild-type and mutant proteins were expressed in Escherichia coli and purified. The mutant and wild-type proteins were characterized by SDS-polyacrylamide gel electrophoresis, Western immunoblotting, gel permeation chromatography, fluorescence, and circular dichroism spectroscopy. Biophysical studies reveal significant differences between the wild-type and mutant proteins. The chaperone-like activity was studied by analyzing the ability of the recombinant proteins to prevent dithiothreitol-induced aggregation of insulin. The mutations R116C in alphaA crystallin and R120G in alphaB crystallin reduce the chaperone-like activity of these proteins significantly. Near UV circular dichroism and intrinsic fluorescence spectra indicate a change in tertiary structure of the mutants. Far UV circular dichroism spectra suggest altered packing of the secondary structural elements. Gel permeation chromatography reveals polydispersity for both of the mutant proteins. An appreciable increase in the molecular mass of the mutant alphaA crystallin is also observed. However, the change in oligomer size of the alphaB mutant is less significant. These results suggest that the conserved arginine of the alpha-crystallin domain of the small heat shock proteins is essential for their structural integrity and subsequent in vivo function.