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1.
Exp Eye Res ; 229: 109429, 2023 04.
Artigo em Inglês | MEDLINE | ID: mdl-36863431

RESUMO

The macular carotenoids lutein and zeaxanthin are taken up from the bloodstream into the human retina through a selective process, for which the HDL cholesterol receptor scavenger receptor BI (SR-BI) in the cells of retinal pigment epithelium (RPE) is thought to be a key mediator. However, the mechanism of SR-BI-mediated selective uptake of macular carotenoids is still not fully understood. Here, we investigate possible mechanisms using biological assays and cultured HEK293 cells, a cell line without endogenous SR-BI expression. Binding affinities between SR-BI and various carotenoids were measured by surface plasmon resonance (SPR) spectroscopy, which shows that SR-BI cannot bind lutein or zeaxanthin specifically. Overexpression of SR-BI in HEK293 cells results in more lutein and zeaxanthin taken up than ß-carotene, and this effect can be eliminated by an SR-BI mutant (C384Y) whose cholesterol uptake tunnel is blocked. Next, we determined the effects of HDL and hepatic lipase (LIPC), SR-BI's partners in HDL cholesterol transport, on SR-BI-mediated carotenoid uptake. HDL addition dramatically reduced lutein, zeaxanthin, and ß-carotene in HEK293 cells expressing SR-BI, but the cellular lutein and zeaxanthin are higher than ß-carotene. LIPC addition increases the uptake of all three carotenoids in HDL-treated cells, and promotes the transport of lutein and zeaxanthin better than ß-carotene. Our results suggest that SR-BI and its HDL cholesterol partner HDL and LIPC may be involved in the selective uptake of macular carotenoids.


Assuntos
Carotenoides , Luteína , Humanos , beta Caroteno , Carotenoides/metabolismo , Antígenos CD36 , Colesterol , HDL-Colesterol/metabolismo , Células HEK293 , Luteína/farmacologia , Receptores Depuradores/metabolismo , Receptores Depuradores Classe B/genética , Receptores Depuradores Classe B/metabolismo , Zeaxantinas
2.
Arch Biochem Biophys ; 716: 109111, 2022 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-34942193

RESUMO

Supplementation with antioxidant carotenoids is a therapeutic strategy to protect against age-related macular degeneration (AMD); however, the transport mechanism of carotenoids from the liver to the retina is still not fully understood. Here, we investigate if HDL serves as the primary transporter for the macular carotenoids. ApoA-I, the key apolipoprotein of HDL, was genetically deleted from BCO2 knockout (Bco2-/-) mice, a macular pigment mouse model capable of accumulating carotenoids in the retina. We then conducted a feeding experiment with a mixed carotenoid chow (lutein:zeaxanthin:ß-carotene = 1:1:1) for one month. HPLC data demonstrated that the total carotenoids were increased in the livers but decreased in the serum, retinal pigment epithelium (RPE)/choroids, and retinas of ApoA-I-/-/Bco2-/- mice compared to Bco2-/- mice. In detail, ApoA-I deficiency caused a significant increase of ß-carotene but not lutein and zeaxanthin in the liver, decreased all three carotenoids in the serum, blocked the majority of zeaxanthin and ß-carotene transport to the RPE/choroid, and dramatically reduced ß-carotene and zeaxanthin but not lutein in the retina. Furthermore, surface plasmon resonance spectroscopy (SPR) data showed that the binding affinity between ApoA-I and ß-carotene â‰« zeaxanthin > lutein. Our results show that carotenoids are transported from the liver to the eye mainly by HDL, and ApoA-I may be involved in the selective delivery of macular carotenoids to the RPE.


Assuntos
Apolipoproteína A-I/genética , Carotenoides/metabolismo , Dioxigenases/genética , Lipoproteínas HDL2/metabolismo , Epitélio Pigmentado da Retina/metabolismo , Animais , Carotenoides/sangue , Modelos Animais de Doenças , Humanos , Fígado , Luteína/metabolismo , Degeneração Macular/metabolismo , Camundongos , Camundongos Knockout , Retina , Zeaxantinas/metabolismo , beta Caroteno/metabolismo
3.
Glycobiology ; 30(7): 433-445, 2020 07 20.
Artigo em Inglês | MEDLINE | ID: mdl-31897472

RESUMO

Morquio syndrome type A, also known as MPS IVA, is a rare autosomal recessive disorder caused by deficiency of N-acetylgalactosamine-6-sulfatase, a lysosomal hydrolase critical in the degradation of keratan sulfate (KS) and chondroitin sulfate (CS). The CS that accumulates in MPS IVA patients has a disease-specific nonreducing end (NRE) terminating with N-acetyl-D-galactosamine 6-sulfate, which can be specifically quantified after enzymatic depolymerization of CS polysaccharide chains. The abundance of N-acetyl-D-galactosamine 6-sulfate over other possible NRE structures is diagnostic for MPS IVA. Here, we describe an assay for the liberation and measurement of N-acetyl-D-galactosamine 6-sulfate and explore its application to MPS IVA patient samples in pilot studies examining disease detection, effects of age and treatment with enzyme-replacement therapy. This assay complements the existing urinary KS assay by quantifying CS-derived substrates, which represent a distinct biochemical aspect of MPS IVA. A more complete understanding of the disease could help to more definitively detect disease across age ranges and more completely measure the pharmacodynamic efficacy of therapies. Larger studies will be needed to clarify the potential value of this CS-derived substrate to manage disease in MPS IVA patients.


Assuntos
Sulfatos de Condroitina/metabolismo , Mucopolissacaridose IV/metabolismo , Adulto , Células Cultivadas , Criança , Sulfatos de Condroitina/química , Sulfatos de Condroitina/urina , Condroitina Sulfatases/metabolismo , Terapia de Reposição de Enzimas , Humanos , Mucopolissacaridose IV/terapia , Mucopolissacaridose IV/urina
4.
Arch Biochem Biophys ; 649: 22-28, 2018 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-29742455

RESUMO

Carotenoid supplementation can improve human visual performance, but there is still no validated rodent model to test their effects on visual function in laboratory animals. We recently showed that mice deficient in ß-carotene oxygenase 2 (BCO2) and/or ß-carotene oxygenase 1 (BCO1) enzymes can accumulate carotenoids in their retinas, allowing us to investigate the effects of carotenoids on the visual performance of mice. Using OptoMotry, a device to measure visual function in rodents, we examined the effect of zeaxanthin, lutein, and ß-carotene on visual performance of various BCO knockout mice. We then transgenically expressed the human zeaxanthin-binding protein GSTP1 (hGSTP1) in the rods of bco2-/- mice to examine if delivering more zeaxanthin to retina will improve their visual function further. The visual performance of bco2-/- mice fed with zeaxanthin or lutein was significantly improved relative to control mice fed with placebo beadlets. ß-Carotene had no significant effect in bco2-/- mice but modestly improved cone visual function of bco1-/- mice. Expression of hGSTP1 in the rods of bco2-/-mice resulted in a 40% increase of retinal zeaxanthin and further improvement of visual performance. This work demonstrates that these "macular pigment mice" may serve as animal models to study carotenoid function in the retina.


Assuntos
Carotenoides/farmacologia , Alimento Funcional , Retina/efeitos dos fármacos , Visão Ocular/efeitos dos fármacos , Animais , Feminino , Alimento Funcional/análise , Glutationa S-Transferase pi/genética , Humanos , Luteína/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Retina/fisiologia , Zeaxantinas/farmacologia , beta Caroteno/farmacologia , beta-Caroteno 15,15'-Mono-Oxigenase/genética
5.
Arch Biochem Biophys ; 634: 21-28, 2017 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-28947101

RESUMO

Carotenoids are plant pigment molecules that are potent antioxidants. Carotenoids cannot be synthesized de novo; therefore, their dietary intake and transport to various tissues are essential to harness their health benefits. Two of the three scavenger receptor class B (SRB) proteins, SR-B1 and CD36, have been implicated as carotenoid transporters in lower species and in various tissues of higher animals. The function of the third SRB protein, SR-B2, in carotenoid transport is unknown. Using surface plasmon resonance (SPR) analyses, we have determined that all three human SRB proteins are capable of binding the macular xanthophyll carotenoids; lutein, zeaxanthin, and meso-zeaxanthin. By over-expressing human SRB proteins in cells that do not endogenously express SRBs, we have determined that lutein uptake is enhanced in the presence of LDL and is mediated by SR-B1 and CD36. SR-B1, SR-B2, and CD36 were able to take up significant amounts of zeaxanthin as well as meso-zeaxanthin, and uptake was increased in the presence of HDL. Our analyses revealed no apparent differences in protein expression profiles of SRBs in central and peripheral regions of human donor tissues, indicating that carotenoid-binding proteins rather than transporters are likely to mediate selective accumulation of carotenoids into the macula.


Assuntos
Lipoproteínas HDL/metabolismo , Lipoproteínas LDL/metabolismo , Proteínas de Membrana Lisossomal/metabolismo , Macula Lutea/metabolismo , Receptores Depuradores/metabolismo , Receptores Depuradores Classe B/metabolismo , Xantofilas/metabolismo , Sítios de Ligação , Transporte Biológico Ativo/fisiologia , Humanos , Técnicas In Vitro , Especificidade de Órgãos , Ligação Proteica , Distribuição Tecidual
6.
Exp Eye Res ; 159: 123-131, 2017 06.
Artigo em Inglês | MEDLINE | ID: mdl-28286282

RESUMO

Carotenoid supplementation can prevent and reduce the risk of age-related macular degeneration (AMD) and other ocular disease, but until now, there has been no validated and well-characterized mouse model which can be employed to investigate the protective mechanism and relevant metabolism of retinal carotenoids. ß-Carotene oxygenases 1 and 2 (BCO1 and BCO2) are the only two carotenoid cleavage enzymes found in animals. Mutations of the bco2 gene may cause accumulation of xanthophyll carotenoids in animal tissues, and BCO1 is involved in regulation of the intestinal absorption of carotenoids. To determine whether or not mice deficient in BCO1 and/or BCO2 can serve as a macular pigment mouse model, we investigated the retinal accumulation of carotenoids in these mice when fed with zeaxanthin, lutein, or ß-carotene using an optimized carotenoid feeding method. HPLC analysis revealed that all three carotenoids were detected in sera, livers, retinal pigment epithelium (RPE)/choroids, and retinas of all of the mice, except that no carotenoid was detectable in the retinas of wild type (WT) mice. Significantly higher amounts of zeaxanthin and lutein accumulated in the retinas of BCO2 knockout (bco2-/-) mice and BCO1/BCO2 double knockout (bco1-/-/bco2-/-) mice relative to BCO1 knockout (bco1-/-) mice, while bco1-/- mice preferred to take up ß-carotene. The levels of zeaxanthin and lutein were higher than ß-carotene levels in the bco1-/-/bco2-/- retina, consistent with preferential uptake of xanthophyll carotenoids by retina. Oxidative metabolites were detected in mice fed with lutein or zeaxanthin but not in mice fed with ß-carotene. These results indicate that bco2-/- and bco1-/-/bco2-/- mice could serve as reasonable non-primate models for macular pigment function in the vertebrate eye, while bco1-/- mice may be more useful for studies related to ß-carotene.


Assuntos
Luteína/metabolismo , Degeneração Macular/metabolismo , Retina/metabolismo , beta Caroteno/metabolismo , Animais , Cromatografia Líquida de Alta Pressão , Modelos Animais de Doenças , Degeneração Macular/patologia , Camundongos , Camundongos Knockout , Oxirredução , Zeaxantinas/metabolismo
7.
Proc Natl Acad Sci U S A ; 111(28): 10173-8, 2014 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-24982131

RESUMO

The macula of the primate retina uniquely concentrates high amounts of the xanthophyll carotenoids lutein, zeaxanthin, and meso-zeaxanthin, but the underlying biochemical mechanisms for this spatial- and species-specific localization have not been fully elucidated. For example, despite abundant retinal levels in mice and primates of a binding protein for zeaxanthin and meso-zeaxanthin, the pi isoform of glutathione S-transferase (GSTP1), only human and monkey retinas naturally contain detectable levels of these carotenoids. We therefore investigated whether or not differences in expression, localization, and activity between mouse and primate carotenoid metabolic enzymes could account for this species-specific difference in retinal accumulation. We focused on ß,ß-carotene-9',10'-dioxygenase (BCO2, also known as BCDO2), the only known mammalian xanthophyll cleavage enzyme. RT-PCR, Western blot analysis, and immunohistochemistry (IHC) confirmed that BCO2 is expressed in both mouse and primate retinas. Cotransfection of expression plasmids of human or mouse BCO2 into Escherichia coli strains engineered to produce zeaxanthin demonstrated that only mouse BCO2 is an active zeaxanthin cleavage enzyme. Surface plasmon resonance (SPR) binding studies showed that the binding affinities between human BCO2 and lutein, zeaxanthin, and meso-zeaxanthin are 10- to 40-fold weaker than those for mouse BCO2, implying that ineffective capture of carotenoids by human BCO2 prevents cleavage of xanthophyll carotenoids. Moreover, BCO2 knockout mice, unlike WT mice, accumulate zeaxanthin in their retinas. Our results provide a novel explanation for how primates uniquely concentrate xanthophyll carotenoids at high levels in retinal tissue.


Assuntos
Dioxigenases/metabolismo , Proteínas do Olho/metabolismo , Luteína/metabolismo , Retina/enzimologia , Xantofilas/metabolismo , Animais , Dioxigenases/genética , Proteínas do Olho/genética , Glutationa S-Transferase pi/genética , Glutationa S-Transferase pi/metabolismo , Humanos , Luteína/genética , Camundongos , Camundongos Knockout , Retina/citologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Especificidade da Espécie , Xantofilas/genética , Zeaxantinas
8.
Arch Biochem Biophys ; 572: 66-72, 2015 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-25513962

RESUMO

The surface plasmon resonance (SPR) biosensor method is a highly sensitive, label-free technique to study the non-covalent interactions of biomolecules, especially protein-protein and protein-small molecule interactions. We have explored this robust biosensor platform to study the interactions of carotenoid-binding proteins and their carotenoid ligands to assess the specificity of interaction, kinetics, affinity, and stoichiometry. These characterizations are important to further study uptake and transport of carotenoids to targeted tissues such as the macula of the human eye. In this review, we present an overview of the SPR method and optimization of assay conditions, and we discuss the particular challenges in studying carotenoid-protein interactions using SPR.


Assuntos
Carotenoides/metabolismo , Proteínas de Transporte/metabolismo , Ressonância de Plasmônio de Superfície/métodos , Transporte Biológico , Humanos , Macula Lutea/metabolismo , Ligação Proteica
9.
Arch Biochem Biophys ; 572: 58-65, 2015 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-25527162

RESUMO

Xanthophyll carotenoids zeaxanthin and lutein play a special role in the prevention and treatment of visual diseases. These carotenoids are not produced by the human body and must be consumed in the diet. On the other hand, extremely low water solubility of these carotenoids and their instability restrict their practical application as components of food or medicinal formulations. Preparation of supramolecular complexes of zeaxanthin and lutein with glycyrrhizic acid, its disodium salt and the natural polysaccharide arabinogalactan allows one to minimize the aforementioned disadvantages when carotenoids are used in food processing as well as for production of therapeutic formulations with enhanced solubility and stability. In the present study, the formation of supramolecular complexes was investigated by NMR relaxation, surface plasmon resonance (SPR) and optical absorption techniques. The complexes increase carotenoid solubility more than 1000-fold. The kinetics of carotenoid decay in reactions with ozone molecules, hydroperoxyl radicals and metal ions were measured in water and organic solutions, and significant increases in oxidation stability of lutein and zeaxanthin in arabinogalactan and glycyrrhizin complexes were detected.


Assuntos
Galactanos/química , Luteína/química , Macula Lutea/química , Oligossacarídeos/química , Água/química , Zeaxantinas/química , Química Farmacêutica , Estabilidade de Medicamentos , Ácido Glicirrízico/química , Oxirredução , Solubilidade
10.
Arch Biochem Biophys ; 539(2): 181-6, 2013 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-23876239

RESUMO

Uptake, transport and stabilization of xanthophylls in the human retina are important components of a complex multistep process that culminates in a non-uniform distribution of these important nutrients in the retina. The process is far from understood; here, we consider the potential role of interphotoreceptor retinoid-binding protein (IRBP) in this process. IRBP is thought to facilitate the exchange of 11-cis-retinal, 11-cis-retinol and all-trans-retinol between the retinal pigment epithelium (RPE), photoreceptors and Müller cells in the visual cycle. Structural and biochemical studies suggest that IRBP has a variety of nonequivalent ligand binding sites that function in this process. IRBP is multifunctional, being able to bind a variety of physiologically significant molecules including fatty acids in the subretinal space. This wide range of binding activities is of particular interest because it is unknown whether the lutein and zeaxanthin found in the macula originate from the choroidal or retinal circulations. If from the choroidal circulation, then IRBP is a likely mediator for their transport across the interphotoreceptor matrix. In this report, we explore the binding interactions of retinoids, fatty acids, and carotenoids with IRBP using surface plasmon resonance (SPR)-based biosensors. IRBP showed similar affinity toward retinoids and carotenoids (1-2 µM), while fatty acids had approximately 10 times less affinity. These results suggest that further studies should be carried out to evaluate whether IRBP has a physiologically relevant role in binding lutein and zeaxanthin in the interphotoreceptor matrix.


Assuntos
Carotenoides/metabolismo , Proteínas do Olho/química , Proteínas do Olho/metabolismo , Proteínas de Ligação ao Retinol/química , Proteínas de Ligação ao Retinol/metabolismo , Ressonância de Plasmônio de Superfície/métodos , Animais , Técnicas Biossensoriais/métodos , Carotenoides/química , Bovinos , Ácidos Graxos não Esterificados/química , Ácidos Graxos não Esterificados/metabolismo , Humanos , Ligantes , Ligação Proteica/fisiologia
11.
Arch Biochem Biophys ; 519(1): 32-7, 2012 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-22286029

RESUMO

The xanthophyll carotenoids lutein and zeaxanthin constitute the major carotenoids of the macular pigment in the human retina where they are thought to act in part to prevent light induced oxidative damage associated with age-related macular degeneration (AMD). The highly selective uptake of these pigments is mediated by specific carotenoid-binding proteins (GSTP1 and StARD3) recently identified in our laboratory. Carotenoids are hydrophobic in nature, so we first systematically optimized carotenoid preparations that are nano-dispersed in aqueous buffers, and then we used a new-generation surface plasmon resonance (SPR) protocol called FastStep™, which is significantly faster than conventional SPR assays. We have explored carotenoid-binding interactions of five proteins: human serum albumin (HSA), ß-lactoglobulin (LG), steroidogenic acute regulatory domain proteins (StARD1, StARD3) and glutathione S- transferase Pi isoform (GSTP1). HSA and LG showed relatively weak interaction with carotenoids (K(D)>1 µM). GSTP1 evidenced high affinity and specificity towards zeaxanthin and meso-zeaxanthin with K(D) values 0.14±0.02 µM and 0.17±0.02 µM, respectively. StARD3 expressed a relative high specificity towards lutein with a K(D) value of 0.59±0.03 µM, whereas StARD1 exhibited a relatively low selectivity and affinity (K(D)>1 µM) towards the various carotenoids tested.


Assuntos
Proteínas de Transporte/metabolismo , Proteínas de Membrana/metabolismo , Fosfoproteínas/metabolismo , Xantofilas/metabolismo , Transporte Biológico/fisiologia , Proteínas de Transporte/química , Proteínas de Transporte/genética , Glutationa S-Transferase pi/química , Glutationa S-Transferase pi/genética , Glutationa S-Transferase pi/metabolismo , Humanos , Proteínas Imobilizadas/química , Proteínas Imobilizadas/genética , Proteínas Imobilizadas/metabolismo , Cinética , Lactoglobulinas/química , Lactoglobulinas/genética , Lactoglobulinas/metabolismo , Luteína/metabolismo , Proteínas de Membrana/química , Proteínas de Membrana/genética , Fosfoproteínas/química , Fosfoproteínas/genética , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Retina/metabolismo , Albumina Sérica/química , Albumina Sérica/genética , Albumina Sérica/metabolismo , Ressonância de Plasmônio de Superfície/métodos , Zeaxantinas
12.
Biochemistry ; 50(13): 2541-9, 2011 Apr 05.
Artigo em Inglês | MEDLINE | ID: mdl-21322544

RESUMO

Lutein, zeaxanthin, and their metabolites are the xanthophyll carotenoids that form the macular pigment of the human retina. Epidemiological evidence suggests that high levels of these carotenoids in the diet, serum, and macula are associated with a decreased risk of age-related macular degeneration (AMD), and the AREDS2 study is prospectively testing this hypothesis. Understanding the biochemical mechanisms underlying the selective uptakes of lutein and zeaxanthin into the human macula may provide important insights into the physiology of the human macula in health and disease. GSTP1 is the macular zeaxanthin-binding protein, but the identity of the human macular lutein-binding protein has remained elusive. Prior identification of the silkworm lutein-binding protein (CBP) as a member of the steroidogenic acute regulatory domain (StARD) protein family and selective labeling of monkey photoreceptor inner segments with an anti-CBP antibody provided an important clue for identifying the primate retina lutein-binding protein. The homology of CBP with all 15 human StARD proteins was analyzed using database searches, Western blotting, and immunohistochemistry, and we here provide evidence to identify StARD3 (also known as MLN64) as a human retinal lutein-binding protein. Antibody to StARD3, N-62 StAR, localizes to all neurons of monkey macular retina and especially cone inner segments and axons, but does not colocalize with the Müller cell marker, glutamine synthetase. Further, recombinant StARD3 selectively binds lutein with high affinity (K(D) = 0.45 µM) when assessed by surface plasmon resonance (SPR) binding assays. Our results demonstrate previously unrecognized, specific interactions of StARD3 with lutein and provide novel avenues for exploring its roles in human macular physiology and disease.


Assuntos
Proteínas de Transporte/metabolismo , Luteína/metabolismo , Macula Lutea/metabolismo , Proteínas de Membrana/metabolismo , Animais , Proteínas de Transporte/química , Bases de Dados de Proteínas , Feminino , Técnica Direta de Fluorescência para Anticorpo , Humanos , Cinética , Macaca mulatta , Macula Lutea/citologia , Masculino , Proteínas de Membrana/química , Camundongos , Proteínas do Tecido Nervoso/química , Proteínas do Tecido Nervoso/metabolismo , Neurônios/metabolismo , Especificidade de Órgãos , RNA Mensageiro/metabolismo , Proteínas Recombinantes/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Homologia de Sequência de Aminoácidos , Ressonância de Plasmônio de Superfície
13.
Photochem Photobiol Sci ; 9(11): 1418-25, 2010 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-20820671

RESUMO

Two dietary carotenoids, lutein and zeaxanthin, are specifically delivered to the human macula at the highest concentration anywhere in the body. Whenever a tissue exhibits highly selective uptake of a compound, it is likely that one or more specific binding proteins are involved in the process. Over the past decade, our laboratory has identified and characterized several carotenoid-binding proteins from human retina including a pi isoform of glutathione S-transferase (GSTP1) as a zeaxanthin-binding protein, a member of the steroidogenic acute regulatory domain (StARD) family as a lutein-binding protein, and tubulin as a less specific, but higher capacity site for carotenoid deposition. In this article, we review the purification and characterization of these carotenoid-binding proteins, and we relate these ocular carotenoid-binding proteins to the transport and uptake role of serum lipoproteins and scavenger receptor proteins in a proposed pathway for macular pigment carotenoid delivery to the human retina.


Assuntos
Carotenoides/metabolismo , Glutationa S-Transferase pi/isolamento & purificação , Proteínas de Membrana Transportadoras/isolamento & purificação , Retina/metabolismo , Transporte Biológico , Glutationa S-Transferase pi/metabolismo , Humanos , Lipoproteínas/metabolismo , Proteínas de Membrana Transportadoras/metabolismo , Ligação Proteica
14.
Biosensors (Basel) ; 6(1)2016 Feb 25.
Artigo em Inglês | MEDLINE | ID: mdl-26927197

RESUMO

Flavonoids are common polyphenolic compounds widely distributed in fruits and vegetables. These pigments have important pharmacological relevance because emerging research suggests possible anti-cancer and anti-inflammatory properties as well other beneficial health effects. These compounds are relatively hydrophobic molecules, suggesting the role of blood transport proteins in their delivery to tissues. In this study, we assess the binding interactions of four flavonoids (kaempferol, luteolin, quercetin, and resveratrol) with human serum albumin (HSA), the most abundant protein in the blood, and with glutathione S-transferase pi isoform-1 (GSTP1), an enzyme with well-characterized hydrophobic binding sites that plays an important role in detoxification of xenobiotics with reduced glutathione, using a novel Taylor dispersion surface plasmon resonance (SPR) technique. For the first time, HSA sites revealed a high-affinity binding site for flavonoid interactions. Out of the four flavonoids that we examined, quercetin and kaempferol showed the strongest equilibrium binding affinities (K(D)) of 63 ± 0.03 nM and 37 ± 0.07 nM, respectively. GSTP1 displayed lower affinities in the micromolar range towards all of the flavonoids tested. The interactions of flavonoids with HSA and GSTP1 were studied successfully using this novel SPR assay method. The new method is compatible with both kinetic and equilibrium analyses.


Assuntos
Flavonoides/metabolismo , Glutationa S-Transferase pi/química , Glutationa S-Transferase pi/metabolismo , Albumina Sérica/química , Albumina Sérica/metabolismo , Sítios de Ligação , Flavonoides/química , Humanos , Quempferóis/metabolismo , Cinética , Luteolina/metabolismo , Ligação Proteica , Quercetina/metabolismo , Resveratrol , Estilbenos/metabolismo , Ressonância de Plasmônio de Superfície
15.
Invest Ophthalmol Vis Sci ; 57(4): 1853-61, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-27082300

RESUMO

PURPOSE: meso-Zeaxanthin is a carotenoid that is rarely encountered in nature outside of the vertebrate eye. It is not a constituent of a normal human diet, yet this carotenoid comprises one-third of the primate macular pigment. In the current study, we undertook a systematic approach to biochemically characterize the production of meso-zeaxanthin in the vertebrate eye. METHODS: Fertilized White Leghorn chicken eggs were analyzed for the presence of carotenoids during development. Yolk, liver, brain, serum, retina, and RPE/choroid were isolated, and carotenoids were extracted. The samples were analyzed on C-30 or chiral HPLC columns to determine the carotenoid composition. RESULTS: Lutein and zeaxanthin were found in all studied nonocular tissues, but no meso-zeaxanthin was ever detected. Among the ocular tissues, the presence of meso-zeaxanthin was consistently observed starting at embryonic day 17 (E17) in the RPE/choroid, several days before its consistent detection in the retina. If RPE/choroid of an embryo was devoid of meso-zeaxanthin, the corresponding retina was always negative as well. CONCLUSIONS: This is the first report of developmentally regulated synthesis of meso-zeaxanthin in a vertebrate system. Our observations suggest that the RPE/choroid is the primary site of meso-zeaxanthin synthesis. Identification of meso-zeaxanthin isomerase enzyme in the developing chicken embryo will facilitate our ability to determine the biochemical mechanisms responsible for production of this unique carotenoid in other higher vertebrates, such as humans.


Assuntos
Corioide/metabolismo , Epitélio Pigmentado da Retina/metabolismo , Animais , Embrião de Galinha , Corioide/embriologia , Cromatografia Líquida de Alta Pressão , Epitélio Pigmentado da Retina/embriologia , Zeaxantinas/biossíntese
16.
Prog Retin Eye Res ; 50: 34-66, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26541886

RESUMO

The human macula uniquely concentrates three carotenoids: lutein, zeaxanthin, and meso-zeaxanthin. Lutein and zeaxanthin must be obtained from dietary sources such as green leafy vegetables and orange and yellow fruits and vegetables, while meso-zeaxanthin is rarely found in diet and is believed to be formed at the macula by metabolic transformations of ingested carotenoids. Epidemiological studies and large-scale clinical trials such as AREDS2 have brought attention to the potential ocular health and functional benefits of these three xanthophyll carotenoids consumed through the diet or supplements, but the basic science and clinical research underlying recommendations for nutritional interventions against age-related macular degeneration and other eye diseases are underappreciated by clinicians and vision researchers alike. In this review article, we first examine the chemistry, biochemistry, biophysics, and physiology of these yellow pigments that are specifically concentrated in the macula lutea through the means of high-affinity binding proteins and specialized transport and metabolic proteins where they play important roles as short-wavelength (blue) light-absorbers and localized, efficient antioxidants in a region at high risk for light-induced oxidative stress. Next, we turn to clinical evidence supporting functional benefits of these carotenoids in normal eyes and for their potential protective actions against ocular disease from infancy to old age.


Assuntos
Oftalmopatias/prevenção & controle , Luteína/fisiologia , Macula Lutea/metabolismo , Zeaxantinas/fisiologia , Animais , Antioxidantes/fisiologia , Dieta , Oftalmopatias/etiologia , Haplorrinos , Humanos , Luteína/administração & dosagem , Luteína/química , Degeneração Macular/metabolismo , Pigmentos da Retina/metabolismo , Zeaxantinas/administração & dosagem , Zeaxantinas/química , Zeaxantinas/metabolismo
17.
Methods Mol Biol ; 898: 41-59, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22711116

RESUMO

Carotenoids are among the most widely distributed pigments in nature, and they are exclusively synthesized by plants and microorganisms. These compounds may serve a protective role against many chronic diseases such as cancers, age-related macular degeneration, and cardiovascular diseases and also act as an excellent antioxidant system within cells. Recent advances in the microbial genome sequences and increased understanding about the genes involved in the carotenoid biosynthetic pathways will assist industrial microbiologists in their exploration of novel microbial carotenoid production strategies. Here we present an overview of microbial carotenogenesis from biochemical, proteomic, and biotechnological points of view.


Assuntos
Carotenoides , Microbiologia , Animais , Biotecnologia , Carotenoides/biossíntese , Carotenoides/metabolismo , Humanos , Proteômica
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