Cataract produced by tyrosinase and tyrosine systems in rabbitens in vitro.
Ontology highlight
ABSTRACT:
SUBMITTER: Srivastava SK
PROVIDER: S-EPMC1187729 | biostudies-other | 1969 May
REPOSITORIES: biostudies-other
Similar Datasets
Project description:Hydroxylation of L-tyrosine to 3,4-dihydroxyphenylalanine (L-DOPA) by immobilized tyrosinase in the presence of ascorbic acid (AH2), which reduces DOPA-quinone to L-DOPA, is characterized by low reaction yields that are mainly caused by the suicide inactivation of tyrosinase by L-DOPA and AH2. The main aim of this work was to compare processes with native and immobilized tyrosinase to identify the conditions that limit suicide inactivation and produce substrate conversions to L-DOPA of above 50% using HPLC analysis. It was shown that immobilized tyrosinase does not suffer from partitioning and diffusion effects, allowing a direct comparison of the reactions performed with both forms of the enzyme. In typical processes, additional aeration was applied and boron ions to produce the L-DOPA and AH2 complex and hydroxylamine to close the cycle of enzyme active center transformations. It was shown that the commonly used pH 9 buffer increased enzyme stability, with concomitant reduced reactivity of 76%, and that under these conditions, the maximal substrate conversion was approximately 25 (native) to 30% (immobilized enzyme). To increase reaction yield, the pH of the reaction mixture was reduced to 8 and 7, producing L-DOPA yields of approximately 95% (native enzyme) and 70% (immobilized). A three-fold increase in the bound enzyme load achieved 95% conversion in two successive runs, but in the third one, tyrosinase lost its activity due to strong suicide inactivation caused by L-DOPA processing. In this case, the cost of the immobilized enzyme preparation is not overcome by its reuse over time, and native tyrosinase may be more economically feasible for a single use in L-DOPA production. The practical importance of the obtained results is that highly efficient hydroxylation of monophenols by tyrosinase can be obtained by selecting the proper reaction pH and is a compromise between complexation and enzyme reactivity.
| S-EPMC5053437 | biostudies-literature
Project description:The Emory mouse is a well-characterized model for age-onset cataract. The purpose of the present study was to identify differentially expressed genes between pre- and postcataract Emory mouse lenses.Eyes were extracted from Emory mice at 3 weeks (precataract) and 7.5 months (postcataract) of age, and lenses were dissected. Lens RNA was compared for gene expression differences by RT-PCR differential display, and transcripts exhibiting altered levels of gene expression were cloned and identified by sequencing. The levels of two transcripts were further evaluated by RT-PCR in 3-week- and 7.5-month-old lenses and the remainder of the eye. The same transcripts were also measured in lenses from three non-Emory mouse strains (FVB/N, 129Sv, and CD1) ages 4 weeks to 11.5 months.Three transcripts were identified as exhibiting altered levels of gene expression between 3-week- and 7.5-month-old Emory mouse lenses. These encoded alphaA-crystallin (decreased), betaA3/A1-crystallin (decreased), and adhesion-related kinase (ARK) receptor tyrosine kinase (increased). Decreased alphaA-crystallin and increased ARK expression were not detected in lenses isolated from three non-Emory mouse strains of similar age. Increased expression of ARK was not detected between 3-week- and 7.5-month-old Emory mouse eye nonlens tissues.The present data confirm that expression of the alphaA-crystallin gene is decreased in cataract in the Emory mouse lens relative to age-matched control lenses and they provide evidence for cataract- and lens-specific upregulation of the ARK receptor tyrosine kinase in the Emory mouse.
| S-EPMC2957826 | biostudies-literature
Project description:Melanogenesis is responsible for skin pigmentation and the enzymatic browning of foods. Tyrosinases play a major role in melanin synthesis, and many attempts have been made to identify new natural tyrosinase inhibitors, but few have sought to do in microbes. Postbiotics are bioactive compounds produced by the metabolism of probiotics and have been reported to be safe and effective. In this study, we evaluated the tyrosinase inhibitory effects of culture supernatants of probiotics and discovered novel bacterial metabolites that can be used as a potent tyrosinase inhibitor based on metabolomics. Cultures of Bifidobacterium bifidum IDCC 4201 and Lactiplantibacillus plantarum IDCC 3501 showed effective anti-tyrosinase, reduced melanin synthesis, and altered protein expression associated with the melanogenesis pathway. Comparative metabolomics analyses conducted by GC-MS identified metabolites commonly produced by B. bifidum and L. plantarum. Of eight selected metabolites, phenyllactic acid exhibited significant tyrosinase-inhibitory activity. Our findings suggest that applications of probiotic culture supernatants containing high amounts of phenyllactic acid have potential use as anti-melanogenesis agents in food and medicines.
| S-EPMC9772785 | biostudies-literature
Project description:We show that mushroom tyrosinase catalyzes the formation of reactive o-quinones on unstructured, tyrosine-rich sequences such as hemagglutinin (HA) tags (YPYDVPDYA). In the absence of exogenous nucleophiles and at low protein concentrations, the o-quinone decomposes with fragmentation of the HA tag. At higher protein concentrations (>5 mg?mL?¹), crosslinking is observed. Besthorn's reagent intercepts the o-quinone to give a characteristic pink complex that can be observed directly on a denaturing SDS-PAGE gel. Similar labeled species can be formed by using other nucleophiles such as Cy5-hydrazide. These reactions are selective for proteins bearing HA and other unstructured poly-tyrosine-containing tags and can be performed in lysates to create specifically tagged proteins.
| S-EPMC3516906 | biostudies-literature
Project description:TYR, DCT and MITF are three important genes involved in maintaining the mature phenotype and producing melanin; they therefore participate in neural crest cell development into melanocytes. Previous studies have revealed that the Wnt signaling factor lymphoid enhancer-binding factor (LEF-1) can enhance DCT and MITF gene expression. However, whether LEF-1 also affects TYR gene expression remains unclear. In the present study, we found that LEF-1 regulated TYR transcription in vitro. LEF-1 overexpression increased TYR gene promoter activity, whereas LEF-1 knockdown by RNA interference significantly decreased TYR expression. Moreover, the core GTTTGAT sequence (-56 to -50) within the TYR promoter is essential for the effect of LEF-1 on TYR expression, and chromatin immunoprecipitation (ChIP) assay indicated that endogenous LEF-1 interacts with the TYR promoter. In addition, we observed a synergistic transactivation of the TYR promoter by LEF-1 and MITF. These data suggest that Wnt signaling plays an important role in regulating melanocyte development and differentiation.
| S-EPMC4651308 | biostudies-literature
Project description:A relationship between coat color and behavioral characteristics has been reported for numerous species. We previously indicated that particular behavioral traits contributing to the genotype at the agouti locus manifest only when possessing a wild-type allele at the albino (i.e., tyrosinase: Tyr) locus. The present study was performed to investigate tyrosinase expression with marked activity in central nervous systems. The whole brain of male B10 and B10-c mice, a B10 congenic strain of the albino locus from BALB/c, at 8 to 9 weeks of age was removed, and obtained several regions of brain, especially catecholaminergic. Comparatively large amounts of Tyr mRNA and its translation products of approximately 68 kDa were found in the regions obtained, and definitely possessed the enzyme activity for the oxidation of L-tyrosine. The present results indicate the possibility that the amount of catecholamines produced in albino mice is higher than that of colored mice due to the deficit in tyrosinase heritably.
| S-EPMC6389515 | biostudies-literature
Project description:1. Melanosomal tyrosinase was isolated from normal C57B1 mice, and a comparison of the tyrosine-hydroxylation and dopa (3,4-dihydroxyphenylalanine)-oxidation activities of this enzyme was made. 2. The results indicate that in the absence of dopa cofactor, this enzyme is capable of tyrosine hydroxylation, but with very little subsequent dopa oxidation and melanin formation. 3. This mechanism of enzyme action may play an important role in the intracellular regulation of melanin formation. 4. Further, dopa appears to act as a positive allosteric effector for tyrosine hydroxylation by tyrosinase, in addition to its known activity as a hydrogen donor for the reaction.
| S-EPMC1163896 | biostudies-other
Project description:1. Frog epidermis tyrosinase was coupled to Sepharose activated with low concentrations of CNBr. The tetrameric form of the enzyme was linked to the matrix via its subunits. Dissociation of the bound active enzyme with guanidinium chloride yielded an active immobilized dimeric derivative. 2. Immobilized dimeric derivative was able to interact with soluble subunits formed transiently during renaturation. An 85% recovery of the native dopa oxidase specific activity was achieved after hybridization. 3. Fluorescence spectra of different immobilized derivatives suggested that tryptophan residues were involved in the interactions between tyrosinase subunits. 4. It is suggested that the activation of the subunits of tyrosinase involves a conformational change towards a more unfolded state, which favours a reassociation to the dimeric active state.
| S-EPMC1163169 | biostudies-other
Project description:The melanogenesis pathway is characterized by a series of reactions catalyzed by key enzymes, such as tyrosinase (TYR), tyrosinase-related protein 2 (TYRP2), and tyrosinase-related protein 1 (TYRP1), to produce melanin pigment. However, in vitro studies of the catalytic activity were incomplete because of a lack of commercially available enzyme substrates, such as dopachrome. Herein, human recombinant intra-melanosomal domains of key enzymes were produced in Trichoplusia ni (T. ni) larvae and then purified using a combination of chromatography techniques in catalytically active form. Using Michaelis–Menten kinetics, the diphenol oxidase activity of tyrosinase achieved the maximum production of native dopachrome at 10 min of incubation at 37 °C for TYR immobilized to magnetic beads (TYR-MB). The presence of dopachrome was confirmed spectrophotometrically at 475 nm through HPLC analysis and in the TYRP2-catalyzed reaction, yielding 5,6-dihydroxyindole-2-carboxylic acid (DHICA). In the TYRP1-driven oxidation of DHICA, the formation of 5,6-indolequinone-2-carboxylic acid (IQCA) was confirmed at ~560 nm. This is the first in vitro reconstitution of the reactions from the melanogenic pathway based on intra-melanosomal domains. In the future, this approach could be used for quantitative in vitro analysis of the melanin pathway, biochemical effects associated with inherited disease-related mutations, and drug screens.
| S-EPMC9820814 | biostudies-literature
Project description:Cancer represents the main cause of morbidity and mortality worldwide, constituting a serious health problem. In this context, melanoma represents the most aggressive and fatal type of skin cancer, with death rates increasing every year. Scientific efforts have been addressed to the development of inhibitors targeting the tyrosinase enzyme as potential anti-melanoma agents due to the importance of this enzyme in melanogenesis biosynthesis. Coumarin-based compounds have shown potential activity as anti-melanoma agents and tyrosinase inhibitors. In this study, coumarin-based derivatives were designed, synthesized, and experimentally evaluated upon tyrosinase. Compound FN-19, a coumarin-thiosemicarbazone analog, exhibited potent anti-tyrosinase activity, with an IC50 value of 42.16 ± 5.16 µM, being more active than ascorbic acid and kojic acid, both reference inhibitors. The kinetic study showed that FN-19 acts as a mixed inhibitor. Still, for this compound, molecular dynamics (MD) simulations were performed to determine the stability of the complex with tyrosinase, generating RMSD, RMSF, and interaction plots. Additionally, docking studies were performed to elucidate the binding pose at the tyrosinase, suggesting that the hydroxyl group of coumarin derivative performs coordinate bonds (bidentate) with the copper(II) ions at distances ranging from 2.09 to 2.61 Å. Then, MM/PBSA calculations revealed that van der Waals interactions are the most relevant intermolecular forces for complex stabilization. Furthermore, it was observed that FN-19 has a binding energy (ΔEMM) value similar to tropolone, a tyrosinase inhibitor. Therefore, the data obtained in this study will be useful for designing and developing novel coumarin-based analogs targeting the tyrosinase enzyme.
| S-EPMC10048804 | biostudies-literature