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1.
Subcell Biochem ; 94: 383-397, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32189308

RESUMO

Albumin is widely conserved from vertebrates to invertebrates, and nature of mammalian albumins permit them to bind various endogenous ligands and drugs in the blood. It is known that at least two major ligand binding sites are present on the albumin molecule, which are referred to as Site I and Site II. These binding sites are thought to be almost completely conserved among mammals, even though the degree of binding to these sites are different depending on the physical and chemical properties of drugs and differences in the microenvironment in the binding pockets. In addition, the binding sites for medium and long-chain fatty acids are also well conserved among mammals, and it is considered that there are at least seven binding sites, including Site I and Site II. These bindings properties of albumin in the blood are also widely known to be important for transporting drugs and fatty acids to various tissues. It can therefore be concluded that albumin is one of the most important serum proteins for various ligands, and information on human albumin can be very useful in predicting the ligand binding properties of the albumin of other vertebrates.


Assuntos
Ácidos Graxos/metabolismo , Preparações Farmacêuticas/metabolismo , Albumina Sérica/metabolismo , Animais , Sítios de Ligação , Ácidos Graxos/química , Humanos , Preparações Farmacêuticas/química , Ligação Proteica , Albumina Sérica/química
2.
Mutat Res ; 849: 503127, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-32087848

RESUMO

The environmental and food contaminant, benzo[a]pyrene {B[a]P, a polycyclic aromatic hydrocarbon (PAH)}, is classified as a human carcinogen by the International Agency for Research on Cancer. The carcinogenicity of B[a]P is linked to the formation of electrophilic metabolites, namely B[a]P-diol epoxides (BPDEs) occurring as stereoisomers. In this work, we quantified the metabolic formation of BPDE isomers and the genotoxic effect in B[a]P-exposed mice, with an aim to estimate the genotoxic potency of B[a]P per in vivo dose of its most potent metabolite [i.e. (+)-anti-BPDE]. The increase in frequency of micronuclei (fMN) in erythrocytes was measured as a biomarker for genotoxic effect. Covalent adducts to serum albumin (SA) and those to DNA from the BPDEs were analysed using liquid chromatography tandem mass spectrometry (LC-MS/MS), as adducts to histidine (BPDE-His-Pro) and deoxyguanosine (BPDE-dG), respectively. For the first time in animal experiments it was possible to resolve adducts to SA from (+)-anti-, (-)-anti- and (±)-syn-BPDE isomers by LC-MS/MS. The adduct levels in the protein were about 16 fmol/mg SA, which was orders of magnitude lower than that in the nucleic acid, 28 pmol/mg DNA, in mice exposed to 100 mg B[a]P per kg body weight (bw). Using SA adduct levels, the in vivo dose of (+)-anti-BPDE was calculated to be approximately 50 nM·h per mg B[a]P per kg bw. This allowed to make a preliminary estimate of the genotoxic potency as 2‰ fMN per µM·h of (+)-anti-BPDE. This estimate was compared to that from another food toxicant, glycidol, studied with similar methods, which indicated that the BPDE has several orders of magnitude higher genotoxic potency. The demonstrated approach on integrating biomarkers of internal dose of a causative agent and that of genotoxic effect for assessing genotoxic potency, using B[a]P as a model, has a potential for improving cancer risk assessment procedures for PAHs.


Assuntos
Benzo(a)pireno/toxicidade , Carcinógenos/toxicidade , Adutos de DNA/química , Micronúcleos com Defeito Cromossômico/estatística & dados numéricos , Albumina Sérica/química , Animais , Biotransformação , Compostos de Epóxi/química , Compostos de Epóxi/toxicidade , Injeções Intraperitoneais , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Testes para Micronúcleos , Propanóis/toxicidade
3.
J Phys Chem Lett ; 11(3): 1170-1177, 2020 Feb 06.
Artigo em Inglês | MEDLINE | ID: mdl-31967479

RESUMO

Understanding nanoscale protein conformational changes at solid-liquid interfaces is critical for predicting how proteins will impact the performance of biomaterials in vivo. Crowding is an important contributor to conformational stability. Here we apply single-molecule high resolution imaging with photobleaching to directly measure dye-conjugated fibronectin's unfolding in varying conditions of crowding with human serum albumin on aminosilanized glass. Using this approach, we identify serum albumin's crowding mechanism. We find that fibronectin achieves larger degrees of unfolding when not crowded by coadsorbed serum albumin. Serum albumin does not as effectively constrict fibronectin's conformation if it is sequentially, rather than simultaneously, introduced, suggesting that serum albumin's crowding mechanism is dependent on its ability to sterically block fibronectin's unfolding during the process of adsorption. Because fibronectin's conformation is dependent on interfacial macromolecular crowding under in vitro conditions, it is important to consider the role of in vivo crowding on protein activity.


Assuntos
Fibronectinas/química , Albumina Sérica/química , Fibronectinas/metabolismo , Vidro/química , Humanos , Nanotecnologia/métodos , Estabilidade Proteica , Desdobramento de Proteína , Albumina Sérica/metabolismo , Propriedades de Superfície
4.
Chemistry ; 26(17): 3661-3687, 2020 Mar 23.
Artigo em Inglês | MEDLINE | ID: mdl-31709642

RESUMO

The two enantiomers of a compound often have profoundly different biological properties and thus their liability to racemisation in aqueous solutions is an important piece of information. The authors reviewed the available data concerning the process of racemisation in vivo, in the presence of biological molecules (e.g., racemase enzymes, serum albumin, cofactors and derivatives) and under purely chemical but aqueous conditions (acid, base and other aqueous systems). Mechanistic studies are described critically in light of reported kinetic data. The types of experimental measurement that can be used to effectively determine rate constants of racemisation in various conditions are discussed and the data they provide is summarised. The proposed origins of enzymatic racemisation are presented and suggest ways to promote the process that are different from processes taking place in bulk water. Experimental and computational studies that provide understanding and quantitative predictions of racemisation risk are also presented.


Assuntos
Racemases e Epimerases/química , Albumina Sérica/química , Cinética , Estereoisomerismo
5.
J Biochem ; 167(2): 165-171, 2020 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-31598674

RESUMO

Protein S-thiolation is a reversible oxidative modification that serves as an oxidative regulatory mechanism for certain enzymes and binding proteins with reactive cysteine residues. It is generally believed that the thiolation occurs at free sulphydryl group of cysteine residues. Meanwhile, despite the fact that disulphide linkages, serving structural and energetic roles in proteins, are stable and inert to oxidative modification, a recent study shows that the thiolation could also occur at protein disulphide linkages when human serum albumin (HSA) was treated with disulphide molecules, such as cystine and homocystine. A chain reaction mechanism has been proposed for the thiolation at disulphide linkages, in which free cysteine (Cys34) is involved in the reaction with disulphide molecules to form free thiols (cysteine or homocysteine) that further react with protein disulphide linkages to form the thiolated cysteine residues in the protein. This review focuses on the recent finding of this unique chain reaction mechanism of protein thiolation.


Assuntos
Dissulfetos/metabolismo , Albumina Sérica/metabolismo , Compostos de Sulfidrila/metabolismo , Animais , Dissulfetos/química , Humanos , Albumina Sérica/química , Compostos de Sulfidrila/química
6.
Chemosphere ; 239: 124667, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31499299

RESUMO

Exposure to air pollution can have both short-term and long-term effects on health. However, the relationships between specific pollutants and their effects can be obscured by characteristics of both the pollution and the exposed population. One way of elucidating the relationships is to link exposures and internal changes at the level of the individual. To this end, we combined personal exposure monitoring (59 individuals, Oxford Street II crossover study) with mass-spectrometry-based analyses of putative serum albumin adducts (fixed-step selected reaction monitoring). We attempted to infer adducts' identities using data from another, higher-resolution mass spectrometry method, and were able to detect a semi-synthetic standard with both methods. A generalised least squares regression method was used to test for associations between amounts of adducts and pollution measures (ambient concentrations of nitrogen dioxide and particulate matter), and between amounts of adducts and short-term health outcomes (measures of lung health and arterial stiffness). Amounts of some putative adducts (e.g., one with a positive mass shift of ∼143 Da) were associated with exposure to pollution (11 associations), and amounts of other adducts were associated with health outcomes (eight associations). Adducts did not appear to provide a link between exposures and short-term health outcomes.


Assuntos
Poluentes Atmosféricos/análise , Poluição do Ar/análise , Exposição Ambiental/efeitos adversos , Monitoramento Ambiental/métodos , Albumina Sérica/química , Estudos Cross-Over , Feminino , Humanos , Masculino , Dióxido de Nitrogênio/análise , Material Particulado/análise , Análise de Regressão
7.
Toxicol In Vitro ; 62: 104697, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31669365

RESUMO

The potential risk of skin sensitisation, associated with the development of allergic contact dermatitis (ACD), is a consideration in the safety assessment of new ingredients for use in personal care products. Protein haptenation in skin by sensitising chemicals is the molecular initiating event causative of skin sensitisation. Current methods for monitoring skin sensitisation rely on limited reactivity assays, motivating interest in the development of proteomic approaches to characterise the skin haptenome. Increasing our mechanistic understanding of skin sensitisation and ACD using proteomics presents an opportunity to develop non-animal predictive methods and/or risk assessment approaches. Previously, we have used a novel stable isotope labelling approach combined with data independent mass spectrometry (HDMSE) to characterise the haptenome for a number of well-known sensitisers. We have now extended this work by characterising the haptenome of the sensitisers Diphenylcyclopropenone (DPCP) and Ethyl Acrylate (EA) with the model protein Human Serum Albumin (HSA) and the complex lysates of the skin keratinocyte, HaCaT cell line. We show that haptenation in complex nucleophilic models is not random, but a specific, low level and reproducible event. Proteomic analysis extends our understanding of sensitiser reactivity beyond simple reactivity assays and offers a route to monitoring haptenation in living cells.


Assuntos
Dermatite Alérgica de Contato/patologia , Haptenos/química , Imunização , Proteínas/química , Proteômica/métodos , Pele/efeitos dos fármacos , Acrilatos/toxicidade , Linhagem Celular , Ciclopropanos/toxicidade , Dermatite Alérgica de Contato/imunologia , Humanos , Espectrometria de Massas , Modelos Moleculares , Albumina Sérica/química
8.
Sensors (Basel) ; 19(24)2019 Dec 09.
Artigo em Inglês | MEDLINE | ID: mdl-31818011

RESUMO

The study describes development of a glycan biosensor for detection of a tumor-associated antibody. The glycan biosensor is built on an electrochemically activated/oxidized graphene screen-printed electrode (GSPE). Oxygen functionalities were subsequently applied for covalent immobilization of human serum albumin (HSA) as a natural nanoscaffold for covalent immobilization of Thomsen-nouvelle (Tn) antigen (GalNAc-O-Ser/Thr) to be fully available for affinity interaction with its analyte-a tumor-associated antibody. The step by step building process of glycan biosensor development was comprehensively characterized using a battery of techniques (scanning electron microscopy, atomic force microscopy, contact angle measurements, secondary ion mass spectrometry, surface plasmon resonance, Raman and energy-dispersive X-ray spectroscopy). Results suggest that electrochemical oxidation of graphene SPE preferentially oxidizes only the surface of graphene flakes within the graphene SPE. Optimization studies revealed the following optimal parameters: activation potential of +1.5 V vs. Ag/AgCl/3 M KCl, activation time of 60 s and concentration of HSA of 0.1 g L-1. Finally, the glycan biosensor was built up able to selectively and sensitively detect its analyte down to low aM concentration. The binding preference of the glycan biosensor was in an agreement with independent surface plasmon resonance analysis.


Assuntos
Anticorpos Antineoplásicos/sangue , Antígenos Glicosídicos Associados a Tumores/química , Técnicas Biossensoriais/métodos , Grafite/química , Anticorpos Antineoplásicos/imunologia , Antígenos Glicosídicos Associados a Tumores/imunologia , Técnicas Eletroquímicas , Eletrodos , Humanos , Limite de Detecção , Albumina Sérica/química
9.
Int J Mol Sci ; 20(24)2019 Dec 12.
Artigo em Inglês | MEDLINE | ID: mdl-31842251

RESUMO

Lateral flow immunochromatographic assays are a powerful diagnostic tool for point-of-care tests, based on their simplicity, specificity, and sensitivity. In this study, a rapid and sensitive gold nanoparticle (AuNP) immunochromatographic strip is produced for detecting aflatoxin B1 (AFB1) in suspicious fungi-contaminated food samples. The 10 nm AuNPs were encompassed by bovine serum albumin (BSA) and AFB1 antibody. Thin-layer chromatography, gel electrophoresis and nuclear magnetic resonance spectroscopy were employed for analysing the chemical complexes. Various concentrations of AFB1 antigen (0-16 ng/mL) were tested with AFB1 antibody-BSA-AuNPs (conjugated AuNPs) and then analysed by scanning electron microscopy, ultraviolet-visible spectroscopy, and Zetasizer. The results showed that the AFB1 antibody was coupled to BSA by the N-hydroxysuccinimide ester method. The AuNPs application has the potential to contribute to AFB1 detection by monitoring a visible colour change from red to purple-blue, with a detection limit of 2 ng/mL in a 96-well plate. The lateral flow immunochromatographic strip tests are rapid, taking less than 10 min., and they have a detection capacity of 10 ng/g. The smartphone analysis of strips provided the results in 3 s, with a detection limit of 0.3 ng/g for AFB1 when the concentration was below 10 ng/g. Excellent agreement was found with AFB1 determination by high-performance liquid chromatography in the determination of AFB1 among 20 samples of peanuts, corn, rice, and bread.


Assuntos
Aflatoxina B1 , Antifúngicos/farmacologia , Ouro , Nanopartículas Metálicas , Fitas Reagentes , Aflatoxina B1/química , Antifúngicos/química , Cromatografia de Afinidade/métodos , Cromatografia Líquida de Alta Pressão , Contaminação de Alimentos/análise , Ouro/química , Nanopartículas Metálicas/química , Estrutura Molecular , Espectroscopia de Prótons por Ressonância Magnética , Albumina Sérica/química
10.
Asian Pac J Cancer Prev ; 20(12): 3603-3609, 2019 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-31870100

RESUMO

BACKGROUND: Ovarian cancer is the seventh most common cancer in females with the highest mortality rate of all gynecological cancers due to its late discovery and ambiguous symptoms. Thus, there is a need for new promising strategies to diagnose ovarian cancer. We aimed at finding a characteristic plasma proteome pattern that could be used for the detection of epithelial ovarian cancer, in comparison with benign ovarian masses and healthy controls. We also aimed at differentiating between profiling of plasma proteins in early and advanced stages of ovarian cancer and between serous and non-serous histopathological types. METHODS: The combination of MagSi-proteomics C8 beads, Ultraflextreme MALDI-TOF and ClinPro Tools software was used to compare the plasma protein spectra from 50 patients with epithelial ovarian cancer, 20 patients with benign ovarian masses and 50 age matched healthy females. RESULTS: A plasma proteome profile of 21 peaks differentiated patients with epithelial ovarian cancer from healthy controls with a sensitivity of 73 % and a specificity of 82.8% upon external validation, while a 5-peak profile differentiated patients with epithelial ovarian cancer from patients with benign ovarian masses with a sensitivity of 81% and a specificity of 73.7%. A 20 peak profile was generated to discriminate between early and late stages of the disease with 88.3% recognition capability and 70% cross validation. CONCLUSION: MALDI-TOF proteomic profiling represents a promising potential tool for diagnosing epithelial ovarian cancer, discriminating between early and advanced stages and between serous and non- serous types.


Assuntos
Carcinoma Epitelial do Ovário/diagnóstico , Perfilação da Expressão Gênica/métodos , Neoplasias Ovarianas/diagnóstico , Proteoma/análise , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Proteínas Sanguíneas/análise , Carcinoma Epitelial do Ovário/patologia , Egito , Feminino , Humanos , Separação Imunomagnética/métodos , Neoplasias Ovarianas/patologia , Albumina Sérica/química
11.
ACS Appl Mater Interfaces ; 11(48): 44989-44998, 2019 Dec 04.
Artigo em Inglês | MEDLINE | ID: mdl-31755268

RESUMO

Tumor hypoxia severely limits the therapeutic efficacy of solid tumors in photodynamic therapy. One strategy is to develop photosensitizers with simultaneously high efficiency in photodynamic (PDT) and photothermal therapies (PTT) in a single natural-origin phototheranostic agent to overcome this problem. However, less attention has been paid to the natural-origin phototheranostic agent with high PDT and PTT efficiencies even though they have negligible side effects and are environmentally sustainable in comparison with many reported phototheranostic agents. In addition, almost all clinical applied photosensitizers are of natural origin so far. Herein, we synthesized a natural product-based hypocrellin derivative (AETHB), with a high singlet oxygen quantum yield of 0.64 as an efficient photosensitizer different from commercially available porphyrin-based photosensitizers. AETHB is further assembled with human serum albumin to construct nanoparticles (HSA-AETHB NPs) with a high photothermal conversion efficiency (more than 50%). As-prepared HSA-AETHB NPs have shown good water solubility and biocompatibility, pH and light stability, wide absorption (400-750 nm), and NIR emission centered at 710 nm. More importantly, HSA-AETHB NPs can be applied for fluorescent/photoacoustic dual-mode imaging and simultaneously highly efficient PDT/PTT in hypoxic solid tumors. Therefore, this natural-origin multifunctional phototheranostic agent is showing very promising for effective, precise, and safe cancer therapy in clinical applications.


Assuntos
Hipertermia Induzida , Neoplasias/terapia , Perileno/análogos & derivados , Fotoquimioterapia , Quinonas/química , Albumina Sérica/química , Animais , Linhagem Celular Tumoral , Feminino , Humanos , Raios Infravermelhos , Camundongos , Camundongos Nus , Nanopartículas/química , Neoplasias/diagnóstico por imagem , Neoplasias/tratamento farmacológico , Perileno/administração & dosagem , Perileno/química , Quinonas/administração & dosagem , Nanomedicina Teranóstica
12.
Chem Biol Interact ; 311: 108787, 2019 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-31400341

RESUMO

Antipsychotic drugs interfere with the antioxidant defense system provoking complex and often toxicological effects. Here we examined differences in plasma albumin reduced free thiol (SH) group content and its reactivity as a consequence of clozapine (CLZ) and ziprasidone (ZIP) binding. Chronic administration of CLZ reduced, whereas treatment with ZIP increased albumin-SH content in rats. Regardless of the ratio of stearic acid (SA) bound to protein, in vitro binding of ZIP to human serum albumin (HSA) increased both the SH group level and reactivity. In contrast, the effect of CLZ on HSA-SH reactivity was dependent on HSA to SA molar ratio. CLZ binding was accompanied by an increase in HSA-SH reactivity in samples with normal, but a reduction of its reactivity level with higher SA/HSA ratio, compared to drug-free samples. We demonstrate by steady-state fluorescence quenching studies that an increase in SA binding to HSA is associated with a significant reduction of binding constant for both antipsychotics. In addition, this is the first report of quantitative characterization of ZIP binding to HSA. Our findings suggest that albumin-SH content and reactivity is modulated by ZIP towards an increased antioxidant defense capacity in circulation, as opposed to CLZ, which can contribute to the safer, more effective treatment of schizophrenia.


Assuntos
Clozapina/química , Ácidos Graxos/química , Piperazinas/química , Albumina Sérica/química , Compostos de Sulfidrila/química , Tiazóis/química , Animais , Clozapina/metabolismo , Ácidos Graxos/metabolismo , Humanos , Masculino , Piperazinas/metabolismo , Ligação Proteica , Ratos , Ratos Wistar , Albumina Sérica/metabolismo , Espectrometria de Fluorescência , Espectrofotometria , Compostos de Sulfidrila/análise , Compostos de Sulfidrila/metabolismo , Tiazóis/metabolismo
13.
Analyst ; 144(17): 5261-5270, 2019 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-31364612

RESUMO

In this work, an enrichment approach for the profiling of N-linked glycans was developed by utilizing a highly porous 3D graphene composite fabricated from graphene oxide nanosheets and a phenol-formaldehyde polymer via graphitization and KOH activation. In tailoring the large surface area (ca. 2213 m2 g-1) and 3D-layered mesoporous structure, the 3D graphene composite demonstrated not only high efficiency in glycan enrichment but also the size-exclusion effect against residual protein interference. For a standard protein ovalbumin digest, 26 N-linked glycans were identified with good repeatability, and the detection limit was as low as 0.25 ng µL-1 with the identification of 13 N-linked glycans (S/N > 10). When the mass ratio of the ovalbumin digest to the interfering proteins, i.e., bovine serum albumin and ovalbumin was 1 : 2000 : 2000, 18 N-linked glycans could still be detected with sufficient signal intensities. From a 60 nL minute complex human serum sample, up to 53 N-linked glycans with S/N > 10 were identified after the 3D graphene enrichment, while only 20 N-linked glycans were identified by the porous graphitized carbon material used for comparison. In addition, the application of the 3D graphene composite in profiling the up-regulated and down-regulated N-linked glycans from the real clinical serum samples of ovarian cancer patients confirmed the potential of the 3D graphene composite for analyzing minute and complicated biological samples.


Assuntos
Grafite/química , Nanoestruturas/química , Ovalbumina/química , Polissacarídeos/análise , Albumina Sérica/química , Feminino , Formaldeído/química , Humanos , Neoplasias Ovarianas/sangue , Oxirredução , Fenol/química , Fenóis/química , Polímeros/química , Polissacarídeos/isolamento & purificação , Porosidade , Soro
14.
Sci Adv ; 5(8): eaaw6081, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31453327

RESUMO

Serum albumin (SA) is used as a carrier to deliver cytotoxic agents to tumors via passive targeting. To further improve SA's tumor targeting capacity, we sought to develop an approach to retain SA-drug conjugates within tumors through a combination of passive and active targeting. SA was recombinantly fused with a collagen-binding domain (CBD) of von Willebrand factor to bind within the tumor stroma after extravasation due to tumor vascular permeability. Doxorubicin (Dox) was conjugated to the CBD-SA via a pH-sensitive linker. Dox-CBD-SA treatment significantly suppressed tumor growth compared to both Dox-SA and aldoxorubicin treatment in a mouse model of breast cancer. Dox-CBD-SA efficiently stimulated host antitumor immunity, resulting in the complete eradication of MC38 colon carcinoma when used in combination with anti-PD-1 checkpoint inhibitor. Dox-CBD-SA decreased adverse events compared to aldoxorubicin. Thus, engineered CBD-SA could be a versatile and clinically relevant drug conjugate carrier protein for treatment of solid tumors.


Assuntos
Antineoplásicos/farmacologia , Neoplasias do Colo/tratamento farmacológico , Doxorrubicina/farmacologia , Portadores de Fármacos/química , Albumina Sérica/química , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/química , Linhagem Celular Tumoral , Doxorrubicina/administração & dosagem , Doxorrubicina/análogos & derivados , Doxorrubicina/química , Feminino , Hidrazonas/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Domínios Proteicos , Fator de von Willebrand/química
15.
Photochem Photobiol Sci ; 18(10): 2461-2468, 2019 Oct 09.
Artigo em Inglês | MEDLINE | ID: mdl-31410432

RESUMO

Complexes of photosensitizers with blood proteins play an essential role in their delivery to the cell, as well as in the efficacy of photodynamic therapy. Biscarbocyanine dye non-covalently binds human serum albumin (HSA), the dissociation constant of the dye with albumin being Kd = (1.7 ± 0.1) × 10-5 M. According to time correlated single photon counting (TCSPC) fluorescence lifetime spectroscopy data, two types of complexes with lifetimes of 1.0 ns and 2.5 ns are formed between the dye and HSA. Confocal fluorescence microscopy has unambiguously shown the penetration of biscarbocyanine into endoplasmic reticulum, lysosomes, mitochondria and nuclei of the cells. The dye demonstrates photocytotoxicity towards the colon carcinoma HCT116 cells with IC50 = 0.3 µM. Hydrophobicity of the polymethine chain and the presence of two positive charges on the dye molecule contribute to the effective binding of the dye with HSA and the penetration into cells. These facts allow considering the biscarbocyanine dye as a promising agent for the photodynamic therapy of cancer.


Assuntos
Carbocianinas/química , Corantes Fluorescentes/química , Albumina Sérica/química , Carbocianinas/metabolismo , Carbocianinas/farmacologia , Núcleo Celular/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Retículo Endoplasmático/metabolismo , Corantes Fluorescentes/metabolismo , Células HCT116 , Humanos , Lisossomos/metabolismo , Ligação Proteica , Albumina Sérica/metabolismo
16.
Molecules ; 24(16)2019 Aug 09.
Artigo em Inglês | MEDLINE | ID: mdl-31395814

RESUMO

Antibacterial lysins are promising proteins that are active against both antibiotic-susceptible and antibiotic-resistant bacterial strains. However, a major limitation of antibacterial lysins is their fast elimination from systemic circulation. PEGylation increases the plasma half-life of lysins but renders them inactive. Here we report the construction of a fusion protein of lysostaphin, a potent anti-staphylococcal lysin, and an albumin-binding domain from streptococcal protein G. The resulting fusion protein was less active than the parent enzyme lysostaphin, but it still retained significant antibacterial activity even when bound to serum albumin. The terminal half-life of the fusion protein in rats was five-fold greater than that of lysostaphin (7.4 vs. 1.5 h), and the area under the curve increased more than 115 times. Most importantly, this increase in systemic circulation time compensated for the decrease in activity. The plasma from rats that received an injection of the fusion protein retained bactericidal activity for up to 7 h, while plasma from rats that received plain lysostaphin lacked any detectable activity after 4 h. To the best of our knowledge, this is the first report of an antibacterial lysin with both improved pharmacokinetic parameters and prolonged bactericidal activity in the systemic circulation.


Assuntos
Proteínas de Bactérias , Lisostafina , Proteínas Recombinantes de Fusão , Albumina Sérica/química , Staphylococcus aureus/crescimento & desenvolvimento , Animais , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/farmacocinética , Proteínas de Bactérias/farmacologia , Feminino , Lisostafina/química , Lisostafina/genética , Lisostafina/farmacocinética , Lisostafina/farmacologia , Ratos , Ratos Wistar , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/farmacocinética , Proteínas Recombinantes de Fusão/farmacologia
17.
J Fluoresc ; 29(5): 1113-1123, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31396829

RESUMO

Swertisin (6-glucosyl flavonoid) and spinosin (2″-ß-O-glucopyranosyl swertisin) are two main components from Ziziphi Spinosae Semen, with anti-anxiety and hypnosis effects. The paper aims to compare the differences between the two compounds binding with serum albumins (BSA and HSA). Swertisin and spinosin statically quench intrinsic fluorescence of serum proteins by binding to proteins to form complexes. The fluorescence quenching rates of BSA induced by swertisin or spinosin are faster than those of HSA resulted by swertisin or spinosin, respectively. Each serum protein has only one binding site respectively accessible to the two compounds. Hydrophobic force and hydrogen bond play the important roles during the biding process of swertisin with proteins, but van der Waals force and hydrogen bond are major driving forces for spinosin binding to proteins. Synchronous fluorescence data show that spinosin binds to BSA and HSA and thus changes Tyr and Trp residue microenvironments, and has a greater effect on the latter. Compared with swertisin, spinosin has a stronger effect on the α-helix of proteins. But the distance between swertisin and proteins is slightly closer than spinosin. These findings will contribute to further understand the reaction of Ziziphi Spinosae Semen in the liver phase I oxidation, intestinal hydrolysis and deparaffin metabolism.


Assuntos
Apigenina/análise , Flavonoides/análise , Fluorescência , Albumina Sérica/química , Ziziphus/química , Animais , Bovinos , Humanos , Conformação Molecular , Espectrometria de Fluorescência
18.
Ann N Y Acad Sci ; 1457(1): 128-141, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31407357

RESUMO

Metallacarboranes are anionic boron clusters with high affinity to serum albumin, ability to cross biological membranes, and no apparent toxicity in vitro and in vivo. Thus, conjugation with cobalt bis(1,2-dicarbollide), [COSAN]- , ([3,3'-Co(1,2-C2 B9 H11 )2 ]- ) may improve the properties of therapeutic peptides or proteins at both molecular and systemic levels. Here, we conjugated [COSAN]- with the therapeutic peptide thymosin ß4 (Tß4), which has a pleiotropic activity that results in enhanced healing and regeneration of injured tissues. Using fluorescence quenching of human serum albumin and surface plasmon resonance techniques, we showed that the conjugates have a high affinity to human serum albumin. Using an in vitro wound closure assay, we showed that conjugation with [COSAN]- enhances the activity of Tß4 toward fibroblasts (MSU1.1 cell line). These results indicate an application of metallacarboranes in the development of analogs of various therapeutic peptides/proteins with superior pharmacological properties.


Assuntos
Albuminas/análise , Boranos/química , Membrana Celular/metabolismo , Cobalto/química , Metais/química , Peptídeos/química , Ânions/química , Linhagem Celular , Dicroísmo Circular , Complexos de Coordenação/química , Fibroblastos/metabolismo , Humanos , Cinética , Estrutura Terciária de Proteína , Albumina Sérica/química , Albumina Sérica Humana/química , Espectrometria de Fluorescência , Espectrofotometria Ultravioleta , Ressonância de Plasmônio de Superfície , Timosina/química
19.
Free Radic Res ; 53(8): 892-900, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31357898

RESUMO

Depending on its redox status, albumin is known to exist as two forms: reduced albumin or human mercaptalbumin (HMA); and oxidised albumin or human nonmercaptalbumin (HNA). The ratio of HNA to HMA is reportedly elevated in several diseases. Since lipid mediators, such as eicosanoids and lysophospholipids, are typically bound to albumin, we examined the possible preferences of lipid mediators for HNA or HMA. We observed that DHA-derived and EPA-derived eicosanoids preferred to be bound to HMA, while the levels of lysophospholipid mediators, such as lysophosphatidic acids and sphingosine 1-phosphate, were higher in the HNA fraction. Considering the bioactivities reported in previous basic studies, these results suggest that proatherosclerotic lipid mediators might generally prefer HNA, while antiatherosclerotic ones might prefer HMA. Oxidative stress affects the redox status of albumin, which might modulate the dynamism of lipid mediators. This pathway might be partly involved in the association between oxidation and atherosclerosis.


Assuntos
Eicosanoides/metabolismo , Lisofosfolipídeos/metabolismo , Albumina Sérica/metabolismo , Aterosclerose/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Oxirredução , Ligação Proteica , Albumina Sérica/química , Albumina Sérica/genética , Albumina Sérica Humana/química , Albumina Sérica Humana/metabolismo
20.
Int J Pharm ; 567: 118458, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-31247277

RESUMO

Successful treatment of age-related macular diseases requires an effective controlled drug release system with less invasive route of administration in the eye to reduce the burden of frequent intravitreal injections for patients. In this study, we developed an episcleral implantable device for sustained release of ranibizumab, and evaluated its efficacy on suppression of laser-induced choroidal neovascularization (CNV) in rats. We tested both biodegradable and non-biodegradable sheet-type devices consisting of crosslinked gelatin/chitosan (Gel/CS) and photopolymerized poly(ethyleneglycol) dimethacrylate that incorporated collagen microparticles (PEGDM/COL). In vitro release studies of FITC-labeled albumin showed a constant release from PEGDM/COL sheets compared to Gel/CS sheets. The Gel/CS sheets gradually biodegraded in the sclera during the 24-week implantation; however, the PEGDM/COL sheets did not degrade. FITC-albumin was detected in the retina during 18 weeks implantation in the PEGDM/COL sheet-treated group, and was detected in the Gel/CS sheet-treated group during 6 weeks implantation. CNV was suppressed 18 weeks after application of ranibizumab-loaded PEGDM/COL sheets compared to a placebo PEGDM/COL sheet-treated group, and to the intravitreal ranibizumab-injected group. In conclusion, the PEGDM/COL sheet device suppressed CNV via a transscleral administration route for 18 weeks, indicating that prolonged sustained ranibizumab release could reduce the burden of repeated intravitreal injections.


Assuntos
Inibidores da Angiogênese/administração & dosagem , Neovascularização de Coroide/tratamento farmacológico , Implantes de Medicamento/administração & dosagem , Ranibizumab/administração & dosagem , Inibidores da Angiogênese/química , Animais , Quitosana/administração & dosagem , Quitosana/química , Colágeno/administração & dosagem , Colágeno/química , Implantes de Medicamento/química , Liberação Controlada de Fármacos , Olho/efeitos dos fármacos , Olho/metabolismo , Olho/patologia , Fluoresceína-5-Isotiocianato/administração & dosagem , Fluoresceína-5-Isotiocianato/análogos & derivados , Fluoresceína-5-Isotiocianato/química , Gelatina/administração & dosagem , Gelatina/química , Lasers , Masculino , Metacrilatos/administração & dosagem , Metacrilatos/química , Polietilenoglicóis/administração & dosagem , Polietilenoglicóis/química , Ranibizumab/química , Ratos Sprague-Dawley , Albumina Sérica/administração & dosagem , Albumina Sérica/química
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