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Lens Protein (lens + protein)

Distribution by Scientific Domains

Chemistry	53%
Medical Sciences	15%

Selected Abstracts

Fluorescence Study on the Interaction Between Hypericin and Lens Protein ",-Crystallin"

PHOTOCHEMISTRY & PHOTOBIOLOGY, Issue 4 2009
Tareq Youssef
Hypericin has been reported as a potent photosensitizing agent exhibiting antiviral, antibacterial, antineoplastic activities. Although its photophysics and mode of action are strongly modulated by the binding protein, detailed information about its mechanism of interaction with possible cellular targets, including proteins, is still lacking. Previous in vitro studies demonstrated that hypericin can be uptaken by intact lens and is able to bind to the major lens protein ",-crystallin." In this study, the mechanism of interaction of this potent drug with ,-crystallin was studied using the chemical denaturant guanidine hydrochloride (GdnHCl) and the hydrophobic surface probe, 8-anilino-1-naphthalenesulfonic acid (ANS). Fluorescence measurements showed that the increased exposure of tryptophan resulting from partial unfolding of ,-crystallin incubated with 1.0 mol L,1 of GdnHCl corresponds to the maximum accessibility of hydrophobic sites to ANS at the same GdnHCl concentration. Interestingly at this additional hydrophobicity of the protein, hypericin exhibited its maximum fluorescence intensity. This in vitro study implied that hydrophobic sites of ,-crystallin play a significant role in its interaction with hypericin. The binding between ,-crystallin and hypericin was found to be enhanced by partial perturbation of the protein. [source]

R120G ,B-crystallin promotes the unfolding of reduced ,-lactalbumin and is inherently unstable

FEBS JOURNAL, Issue 3 2005
Teresa M. Treweek
,-Crystallin is the principal lens protein which, in addition to its structural role, also acts as a molecular chaperone, to prevent aggregation and precipitation of other lens proteins. One of its two subunits, ,B-crystallin, is also expressed in many nonlenticular tissues, and a natural missense mutation, R120G, has been associated with cataract and desmin-related myopathy, a disorder of skeletal muscles [Vicart P, Caron A, Guicheney P, Li Z, Prevost MC, Faure A, Chateau D, Chapon F, Tome F, Dupret JM, Paulin D & Fardeau M (1998) Nat Genet20, 92,95]. In the present study, real-time 1H-NMR spectroscopy showed that the ability of R120G ,B-crystallin to stabilize the partially folded, molten globule state of ,-lactalbumin was significantly reduced in comparison with wild-type ,B-crystallin. The mutant showed enhanced interaction with, and promoted unfolding of, reduced ,-lactalbumin, but showed limited chaperone activity for other target proteins. Using NMR spectroscopy, gel electrophoresis, and MS, we observed that, unlike the wild-type protein, R120G ,B-crystallin is intrinsically unstable in solution, with unfolding of the protein over time leading to aggregation and progressive truncation from the C-terminus. Light scattering, MS, and size-exclusion chromatography data indicated that R120G ,B-crystallin exists as a larger oligomer than wild-type ,B-crystallin, and its size increases with time. It is likely that removal of the positive charge from R120 of ,B-crystallin causes partial unfolding, increased exposure of hydrophobic regions, and enhances its susceptibility to proteolysis, thus reducing its solubility and promoting its aggregation and complexation with other proteins. These characteristics may explain the involvement of R120G ,B-crystallin with human disease states. [source]

Detection and Prevention of Ocular Phototoxicity of Ciprofloxacin and Other Fluoroquinolone Antibiotics,

PHOTOCHEMISTRY & PHOTOBIOLOGY, Issue 4 2010
Baozhong Zhao
Fluoroquinolone (FLQ) drugs are a potent family of antibiotics used to treat infections including ocular infections. To determine if these antibiotics may be phototoxic to the eye, we exposed human lens epithelial cells to 0.125,1 mm FLQs (ciprofloxacin [Cipro], lomefloxacin [Lome], norfloxacin [Nor] and ofloxacin [Ofl]), the precursor quinolone nalidixic acid (Nalid) and UVA radiation (2.5 J cm,2). Based on fluorescence confocal microscopy, FLQs are diffused throughout the cytoplasm and preferentially located in the lysosomes of lens epithelial cells. Neither FLQ exposure alone nor UVA exposure alone reduced cell viability. However, with exposure to UVA radiation the FLQs studied (Cipro, Nor, Lome and Ofl) induced a phototoxic reaction that included necrosis, apoptosis, loss of cell viability as measured by MTS, and membrane damage as determined by the lactate dehydrogenase assay. Both Nalid and all FLQs studied (Cipro, Nor, Lome and Ofl) photopolymerized the lens protein ,-crystallin. Phototoxic damage to lens epithelial cells and/or ,-crystallin will lead to a loss of transparency of the human lens. However, if precautions are taken to filter all UV radiation from the eye while taking these antibiotics, eye damage may be prevented. [source]

Fluorescence Study on the Interaction Between Hypericin and Lens Protein ",-Crystallin"

Deamidation destabilizes and triggers aggregation of a lens protein, ,A3-crystallin

PROTEIN SCIENCE, Issue 9 2008
Takumi Takata
Abstract Protein aggregation is a hallmark of several neurodegenerative diseases and also of cataracts. The major proteins in the lens of the eye are crystallins, which accumulate throughout life and are extensively modified. Deamidation is the major modification in the lens during aging and cataracts. Among the crystallins, the ,A3-subunit has been found to have multiple sites of deamidation associated with the insoluble proteins in vivo. Several sites were predicted to be exposed on the surface of ,A3 and were investigated in this study. Deamidation was mimicked by site-directed mutagenesis at Q42 and N54 on the N-terminal domain, N133 and N155 on the C-terminal domain, and N120 in the peptide connecting the domains. Deamidation altered the tertiary structure without disrupting the secondary structure or the dimer formation of ,A3. Deamidations in the C-terminal domain and in the connecting peptide decreased stability to a greater extent than deamidations in the N-terminal domain. Deamidation at N54 and N155 also disrupted the association with the ,B1-subunit. Sedimentation velocity experiments integrated with high-resolution analysis detected soluble aggregates at 15%,20% in all deamidated proteins, but not in wild-type ,A3. These aggregates had elevated frictional ratios, suggesting that they were elongated. The detection of aggregates in vitro strongly suggests that deamidation may contribute to protein aggregation in the lens. A potential mechanism may include decreased stability and/or altered interactions with other ,-subunits. Understanding the role of deamidation in the long-lived crystallins has important implications in other aggregation diseases. [source]

Interdomain side-chain interactions in human ,D crystallin influencing folding and stability

PROTEIN SCIENCE, Issue 8 2005
Shannon L. Flaugh
Abstract Human ,D crystallin (H,D-Crys) is a two domain, ,-sheet eye lens protein that must remain soluble throughout life for lens transparency. Single amino acid substitutions of H,D-Crys are associated with juvenile-onset cataracts. Features of the interface between the two domains conserved among ,-crystallins are a central six-residue hydrophobic cluster, and two pairs of interacting residues flanking the cluster. In H,D-Crys these pairs are Gln54/Gln143 and Arg79/Met147. We previously reported contributions of the hydrophobic cluster residues to protein stability. In this study alanine substitutions of the flanking residue pairs were constructed and analyzed. Equilibrium unfolding/refolding experiments at 37°C revealed a plateau in the unfolding/refolding transitions, suggesting population of a partially folded intermediate with a folded C-terminal domain (C-td) and unfolded N-terminal domain (N-td). The N-td was destabilized by substituting residues from both domains. In contrast, the C-td was not significantly affected by substitutions of either domain. Refolding rates of the N-td were significantly decreased for mutants of either domain. In contrast, refolding rates of the C-td were similar to wild type for mutants of either domain. Therefore, domain interface residues of the folded C-td probably nucleate refolding of the N-td. We suggest that these residues stabilize the native state by shielding the central hydrophobic cluster from solvent. Glutamine and methionine side chains are among the residues covalently damaged in aged and cataractous lenses. Such damage may generate partially unfolded, aggregation- prone conformations of H,D-Crys that could be significant in cataract. [source]

Contributions of hydrophobic domain interface interactions to the folding and stability of human ,D-crystallin

PROTEIN SCIENCE, Issue 3 2005
Shannon L. Flaugh
Abstract Human ,D-crystallin (H,D-Crys) is a monomeric eye lens protein composed of two highly homologous ,-sheet domains. The domains interact through interdomain side chain contacts forming two structurally distinct regions, a central hydrophobic cluster and peripheral residues. The hydrophobic cluster contains Met43, Phe56, and Ile81 from the N-terminal domain (N-td) and Val132, Leu145, and Val170 from the C-terminal domain (C-td). Equilibrium unfolding/refolding of wild-type H,D-Crys in guanidine hydrochloride (GuHCl) was best fit to a three-state model with transition midpoints of 2.2 and 2.8 M GuHCl. The two transitions likely corresponded to sequential unfolding/refolding of the N-td and the C-td. Previous kinetic experiments revealed that the C-td refolds more rapidly than the N-td. We constructed alanine substitutions of the hydrophobic interface residues to analyze their roles in folding and stability. After purification from E. coli, all mutant proteins adopted a native-like structure similar to wild type. The mutants F56A, I81A, V132A, and L145A had a destabilized N-td, causing greater population of the single folded domain intermediate. Compared to wild type, these mutants also had reduced rates for productive refolding of the N-td but not the C-td. These data suggest a refolding pathway where the domain interface residues of the refolded C-td act as a nucleating center for refolding of the N-td. Specificity of domain interface interactions is likely important for preventing incorrect associations in the high protein concentrations of the lens nucleus. [source]

Novel spliced form of a lens protein as a novel lung cancer antigen, Lengsin splicing variant 4

CANCER SCIENCE, Issue 8 2009
Munehide Nakatsugawa
A glutamine synthetase I family protein, Lengsin, was previously identified as a novel lens-specific transcript in the vertebrate eye. In this report, we show for the first time that Lengsin is a novel tumor-associated antigen expressed ectopically in lung cancer. Interestingly, a novel spliced form of human Lengsin termed ,splicing variant 4', gaining exon 3 that codes extra 63 amino acids, is the dominant transcript form in lung cancer cells. Lengsin mRNA could be detected in 7 of 12 (58%) lung cancer cell lines and 7 of 7 (100%) surgically resected lung cancer tissues. On the other hand, Lengsin transcripts could not be detected in normal major tissues or in other cancer cell lines, including melanoma, colorectal carcinoma, breast carcinoma and hepatocellular carcinoma. In addition, knockdown of Lengsin mRNA with RNAi caused cell death and a decrease of cell viability, suggesting that Lengsin has some essential role in cell survival. Since the lens is an immune-privileged site, we regard Lengsin as a highly immunogenic cancer antigen. Anti-Lengsin autoantibodies were detectable in sera of lung cancer patients, although these patients did not show any lens-related disturbances. Hence, Lengsin splicing variant 4 might be an immunogenic lung cancer-specific antigen that is suitable as a diagnostic marker and for molecular targeting therapy, including immunotherapy. (Cancer Sci 2009) [source]

R120G ,B-crystallin promotes the unfolding of reduced ,-lactalbumin and is inherently unstable

Inhibition of ascorbic acid-induced modifications in lens proteins by peptides

JOURNAL OF PEPTIDE SCIENCE, Issue 3 2003
Mariana Argirova
Abstract The effects of three dipeptides L -phenylalanyl-glycine, glycyl- L -phenylalanine, and aspartame (L -aspartyl- L -phenylalanine, methyl ester) as inhibitors of the ascorbic acid-induced modifications in lens proteins were studied. Their efficiency was compared to that of two known inhibitors,aminoguanidine and carnosine. The tested dipeptides diminished protein carbonyl content by 32,58% and most moderated the formation of chromophores, as measured by the absorbency at 325 nm of the glycated proteins. The appearance of non-tryptophan fluorescence (excitation 340 nm/emission 410 nm) was observed for proteins glycated with ascorbic acid. All of the dipeptides examined, as well as aminoguanidine, decreased this glycation-related fluorescence. The potential inhibitors prevented the intensive formation of very high molecular weight aggregates. A competitive mechanism of their inhibitory effect was proposed, based on the reactivity of individual substances toward ascorbic acid. These findings indicate that they have a potential for use as alternatives for aminoguanidine as an anti-glycation agent. Copyright © 2003 European Peptide Society and John Wiley & Sons, Ltd. [source]

Dietary histidine affects lens protein turnover and synthesis of N-acetylhistidine in Atlantic salmon (Salmo salar L.) undergoing parr,smolt transformation

AQUACULTURE NUTRITION, Issue 5 2005
O. BRECK
Abstract This study was conducted to investigate protein synthesis rates and metabolism of histidine (His)-derivatives in lenses of Atlantic salmon (Salmo salar L.) of different dietary His background during parr,smolt transformation. Two populations of Atlantic salmon parr of equal origin were established in freshwater (FW), 3 months prior to transfer to seawater (SW). The populations were fed either a control diet (CD) containing 8.9 g kg,1 His or the same diet added crystalline His to a total level of 14.2 g kg,1 (HD). On the basis of these two populations, 14C His force-feeding studies were performed; in FW 3 weeks prior to sea transfer and in SW 6 weeks after transfer. The studies were conducted by force-feeding the respective diets enriched with 14C labelled His, with subsequent measurements of incorporation of 14C His into lens free amino acid pool, as well as into lens proteins and other free His pool fractions. The latter included the major lens imidazole N-acetylhistidine (NAH). Lens concentrations of His and NAH were clearly influenced by dietary His history, both in parr and smolt. The lens His and NAH concentrations in the CD population were considerably lower in SW than in FW, while in the HD group the His level was equal and the NAH level 50% higher in SW than in FW. Fractional synthesis rate for NAH, KS (NAH), in FW was 8.2 and 4.2 ,mol g,1 day,1 for fish in the CD and HD populations, respectively. The corresponding KS (NAH) values in SW were 5.1 and 33.0 ,mol g,1 day,1. Our data show that free His is rapidly converted to NAH in the lens, and that NAH seems to have a very high turnover, especially in salmon reared in SW. Fractional synthesis rate for lens proteins, KS (PROTEIN), ranged between 1.8 and 17.3% day,1 (182 and 2791 ,g g,1 day,1, respectively), and was generally higher in SW than in FW (P < 0.01). In SW, KS (PROTEIN) was highest in fish in the HD population (P < 0.05), whereas lens protein retention in the HD group was significantly lower than the CD group (P = 0.01). In a second model assuming that His from lens NAH is available for protein synthesis, calculated values of KS (PROTEIN) ranged between 0.17% day,1 (17.6 ,g g,1 day,1) and 0.48% day,1 (70.2 ,g g,1 day,1). Cataract scores recorded in the His populations at a later point (day 204), showed that the CD fish had significantly higher mean cataract scores than individuals in the HD population (P < 0.01), confirming that low levels of lens His and NAH are associated with cataract development. [source]

Antioxidants reduce diabetic damage in bovine lenses in culture

ACTA OPHTHALMOLOGICA, Issue 2009
A DOVRAT
Purpose Background: There are several theories regarding possible mechanisms leading to diabetic cataract. Few of them include oxidation stress. Aims: Investigation of the mechanisms of cataract formation under diabetic conditions, and examination of the effects of N-acetyl-L-cysteine (NAC), (which is a precursor of glutathione and an anti-inflammatory agent) and derivatives of Desferrioxamine (DFO)(which is an iron chelator and reduces oxidative stress) on diabetic cataract. Methods The experiments included 78 bovine lenses. The lenses were divided into eight different treatments including controls and lenses incubated with high glucose levels (450 mg %) with or without each one of the antioxidants. The intact lenses were incubated for a period of two weeks in our special organ culture conditions. Lens optical quality was analyzed every 24 hours. At the end of the culture period, oxidation was followed in the lens epithelial cells with dichlorofluorescein assay and lens proteins were analyzed by SDS and 2D gel electrophoresis. Results High levels of glucose in the culture medium caused optical damage to bovine lenses, increased lens volume due to swelling, increased oxidation of lens epithelial cells, and caused changes in lens beta crystallin. The anti-oxidants reduced this damage. NAC and Zn-DFO protected the lenses better than DFO. Conclusion Antioxidants can protect the lens from high glucose damage. This study was supported in part by a grant from the Esther and Chaim Coppel Trust and by the Guzik Ophthalmology Research Fund [source]