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1.
Rev. cient. odontol ; 9(1): e051, ene.-mar. 2021. ilus
Artículo en Español | LILACS, LIPECS | ID: biblio-1254403

RESUMEN

La mucopolisacaridosis tipo VI, también conocida como síndrome de Maroteaux-Lamy, es un trastorno lisosómico autosómico recesivo, causado por la deficiencia de la enzima arilsulfatasa B, lo que conduce a la acumulación de dermatán sulfato en los tejidos y su excreción urinaria. La deposición de mucopolisacáridos genera un trastorno progresivo que afecta a múltiples órganos y que, a menudo, resulta en la muerte a temprana edad. Esta enfermedad tiene varias manifestaciones orales, entre las que destacan las complicaciones dentales, que pueden ser graves e incluir folículos similares a quistes dentígeros, maloclusiones, defectos condilares e hiperplasia gingival, además de características clínicas como cuello corto, opacidad corneal, macroglosia y agrandamiento del cráneo, dimensión anteroposterior larga y mano en garra. Se presenta el caso de un paciente de 14 meses de edad que acudió a consulta de odontopediatría por episodios de fiebre, bajo peso e hiperplasia gingival severa. El examen físico evidenció facies tosca, cuello corto, pectus excavatus, manos con disminución en agarre y retardo en el neurodesarrollo. El examen intraoral halló retardo de la erupción dental, hiperplasia gingival generalizada y paladar con poco crecimiento transversal. El examen radiográfico detectó órganos dentarios incluidos y mala posición en el sector anterior, molares superiores dentro del seno maxilar y caninos inferiores rotados. El paciente fue remitido a medicina para exámenes bioquímicos y genéticos para definir el diagnóstico. La bioquímica reveló MPS tipo VI, lo que fue confirmado mediante prueba molecular. Las manifestaciones clínicas en este caso corresponden a la forma clínica de progresión rápida reportada en estos pacientes: talla baja, malformaciones esqueléticas y alteraciones a nivel oral. Los niños con MPS VI grave comienzan temprano y progresan rápidamente, las radiografías óseas y la medición de GAG en orina son útiles para el diagnóstico con actividad de la enzima ARSB y genética. Es necesario fortalecer el conocimiento en odontología y la población en general sobre las características clínicas de mucopolisacáridos tipo VI para tener un diagnóstico temprano y un mejor manejo de patologías en estos pacientes. (AU)


Mucopolysaccharidosis type VI, also known as Maroteaux-Lamy syndrome, is an autosomal recessive lysosomal disorder, due to the deficiency of the enzyme arylsulfatase B that leads to the accumulation of dermatan sulfate in the tissues and its urinary excretion. Mucopolysaccharide deposition leads to a progressive disorder affecting multiple organs that often results in death at a young age. This disease has several oral manifestations, among which dental complications can be serious and include follicles similar to dentigerous cysts, malocclusions, condylar defects and gingival hyperplasia, in addition to a short neck, corneal opacity, macroglossia, skull enlargement, anteroposterior dimension long, claw hand is some of the clinical features. A case of a 14-month-old patient is presented, who attended a pediatric dentistry consultation for episodes of fever, low weight, severe gingival hyperplasia. Physical examination revealed coarse facies, short neck, pectus excavatus, hands with decreased grip, and neurodevelopmental delay. On intraoral examination, dental eruption delayed, generalized gingival hyperplasia, palate with little transverse growth. On radiographic examination, dental organs included and poor position in the anterior sector, upper molars within the maxillary sinus, rotated lower canines. He is referred to medicine for biochemical tests and genetics for diagnosis. Detailed biochemistry MPS type VI, confirmed by molecular testing. The clinical manifestations in this case correspond to the clinical form of rapid progression reported in these patients. They report: short stature, skeletal malformations and alterations at the oral level. Children with severe MPS VI start early and progress rapidly, bone radiographs and urine GAG measurement are helpful for diagnosis with genetic and ARSB enzyme activity. It is necessary to strengthen the knowledge in dentistry and the general population about the clinical characteristics of type VI mucopolysaccharides in order to have an early diagnosis and management of pathologies in these patients. (AU)


Asunto(s)
Humanos , Femenino , Lactante , Arilsulfatasas , Mucopolisacaridosis VI , Dermatán Sulfato , Hiperplasia Gingival , Glicosaminoglicanos
2.
J Pharm Biomed Anal ; 195: 113818, 2021 Feb 20.
Artículo en Inglés | MEDLINE | ID: mdl-33342568

RESUMEN

Sulfation of metabolites is the second highest phase II modification in humans, which plays a critical role in the xenobiotics clearance process and gut microbiota-host co-metabolism. Besides the main function to remove xenobiotics from the body, sulfated metabolites have also been linked to inflammation, bacterial pathogenesis and metabolic disorders. A better understanding of how these metabolites impact the human body has turned into an important research area. Analytical methods for selective identification of this metabolite class are scarce. We have recently developed an assay utilizing the arylsulfatase from Helix pomatia due to a high substrate promiscuity combined with state-of-the-art metabolomics bioinformatic analysis for the selective identification of O-sulfated metabolites in human samples. This enzyme requires a multistep purification process as highest purity is needed for the developed mass spectrometric assay. In this study, we have utilized a new and recombinant overexpressed arylsulfatase (ASPC) for the selective identification of organic sulfate esters in human urine samples. We have compared the substrate conversion in urine samples and substrate specificity of this enzyme with purified arylsulfatase from Helix pomatia. Our analysis of urine samples revealed that both enzymes can be utilized for the selective analysis and discovery of sulfated metabolites with high promiscuity as demonstrated by equal hydrolysis of 108 substrates including sulfated conjugates of 27 metabolites of microbial origin. Importantly, we also identified 21 substrates in human urine samples that are exclusively hydrolyzed by ASPC and application of this enzyme increases the discovery of unknown sulfated metabolites with a higher scaffold diversity.


Asunto(s)
Arilsulfatasas , Microbioma Gastrointestinal , Humanos , Espectrometría de Masas , Metabolómica , Sulfatos
3.
Biochem J ; 477(17): 3433-3451, 2020 09 18.
Artículo en Inglés | MEDLINE | ID: mdl-32856704

RESUMEN

Mucopolysaccharidoses comprise a group of rare metabolic diseases, in which the lysosomal degradation of glycosaminoglycans (GAGs) is impaired due to genetically inherited defects of lysosomal enzymes involved in GAG catabolism. The resulting intralysosomal accumulation of GAG-derived metabolites consequently manifests in neurological symptoms and also peripheral abnormalities in various tissues like liver, kidney, spleen and bone. As each GAG consists of differently sulfated disaccharide units, it needs a specific, but also partly overlapping set of lysosomal enzymes to accomplish their complete degradation. Recently, we identified and characterized the lysosomal enzyme arylsulfatase K (Arsk) exhibiting glucuronate-2-sulfatase activity as needed for the degradation of heparan sulfate (HS), chondroitin sulfate (CS) and dermatan sulfate (DS). In the present study, we investigated the physiological relevance of Arsk by means of a constitutive Arsk knockout mouse model. A complete lack of glucuronate desulfation was demonstrated by a specific enzyme activity assay. Arsk-deficient mice show, in an organ-specific manner, a moderate accumulation of HS and CS metabolites characterized by 2-O-sulfated glucuronate moieties at their non-reducing ends. Pathophysiological studies reflect a rather mild phenotype including behavioral changes. Interestingly, no prominent lysosomal storage pathology like bone abnormalities were detected. Our results from the Arsk mouse model suggest a new although mild form of mucopolysacharidose (MPS), which we designate MPS type IIB.


Asunto(s)
Arilsulfatasas/metabolismo , Sulfatos de Condroitina/metabolismo , Heparitina Sulfato/metabolismo , Mucopolisacaridosis/metabolismo , Animales , Arilsulfatasas/genética , Sulfatos de Condroitina/genética , Activación Enzimática , Heparitina Sulfato/genética , Ratones , Ratones Noqueados , Mucopolisacaridosis/genética
4.
Int J Mol Sci ; 21(14)2020 Jul 12.
Artículo en Inglés | MEDLINE | ID: mdl-32664626

RESUMEN

Perturbations of glycosaminoglycan metabolism lead to mucopolysaccharidoses (MPS)-lysosomal storage diseases. One type of MPS (type VI) is associated with a deficiency of arylsulfatase B (ARSB), for which we previously established a cellular model using pulmonary artery endothelial cells with a silenced ARSB gene. Here, we explored the effects of silencing the ARSB gene on the growth of human pulmonary artery smooth muscle cells in the presence of different concentrations of dermatan sulfate (DS). The viability of pulmonary artery smooth muscle cells with a silenced ARSB gene was stimulated by the dermatan sulfate. In contrast, the growth of pulmonary artery endothelial cells was not affected. As shown by microarray analysis, the expression of the arylsulfatase G (ARSG) in pulmonary artery smooth muscle cells increased after silencing the arylsulfatase B gene, but the expression of genes encoding other enzymes involved in the degradation of dermatan sulfate did not. The active site of arylsulfatase G closely resembles that of arylsulfatase B, as shown by molecular modeling. Together, these results lead us to propose that arylsulfatase G can take part in DS degradation; therefore, it can affect the functioning of the cells with a silenced arylsulfatase B gene.


Asunto(s)
Dermatán Sulfato/metabolismo , Miocitos del Músculo Liso/enzimología , N-Acetilgalactosamina-4-Sulfatasa/fisiología , Secuencia de Aminoácidos , Arilsulfatasas/biosíntesis , Arilsulfatasas/química , Arilsulfatasas/genética , Dominio Catalítico , Dermatán Sulfato/farmacología , Células Endoteliales/efectos de los fármacos , Células Endoteliales/enzimología , Silenciador del Gen , Humanos , Modelos Moleculares , Mucopolisacaridosis VI/metabolismo , Miocitos del Músculo Liso/efectos de los fármacos , N-Acetilgalactosamina-4-Sulfatasa/química , Especificidad de Órganos , Unión Proteica , Conformación Proteica , Arteria Pulmonar/citología , ARN Mensajero/biosíntesis , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Análisis de Matrices Tisulares , Regulación hacia Arriba
5.
ACS Chem Biol ; 15(6): 1349-1357, 2020 06 19.
Artículo en Inglés | MEDLINE | ID: mdl-32239919

RESUMEN

Arylsulfatase A (ARSA) plays a crucial role in the reproduction of mammals due to its involvement in the specific gamete interaction preceding sperm and egg fusion leading to fertilization. Recently, it has been shown that zona pellucida (ZP) sperm binding and in vivo fertilization in mice are markedly hampered by using a specific anti-ARSA antibody. Herein, the design and discovery of the first ARSA small molecule inhibitor based on a coumarin-containing polycycle are presented. Through a structure-based approach applied on our in-house library, compound 1r was identified as an ARSA reversible inhibitor (ARSAi); then its activity was validated through both surface plasmon resonance and biochemical inhibition experiments, the first providing a KD value of 21 µM and the latter an IC50 value of 13.2 µM. Further investigations highlighted that compound 1r induced 20% sperm death at 25 µM and also impaired sperm motility; nevertheless both the effects were mediated by ROS production, since they were rescued by the cotreatment of 1r and N-acetyl cysteine (NAC). Interestingly, while 1r was not able to hamper the ZP/sperm binding, it markedly decreased the in vitro oocyte fertilization by mouse sperm up to 60%. Notably, this effect was not hampered by 1r/NAC coadministration, hence allowing the ruling out of an ROS-dependent mechanism. In conclusion, herein is reported the first ever hit of ARSAi as a chemical tool that will enable better exploration of ARSA's biological role in fertilization as well as provide a starting point for developing 1r structure optimization aimed at increasing enzyme inhibition potency but also providing a deeper understanding of the involvement of ARSA in the fertilization pathway mechanism.


Asunto(s)
Arilsulfatasas/antagonistas & inhibidores , Cumarinas/farmacología , Inhibidores Enzimáticos/farmacología , Fertilización/efectos de los fármacos , Oocitos/efectos de los fármacos , Animales , Arilsulfatasas/metabolismo , Línea Celular Tumoral , Cumarinas/química , Descubrimiento de Drogas , Inhibidores Enzimáticos/química , Femenino , Humanos , Masculino , Ratones , Simulación del Acoplamiento Molecular , Oocitos/fisiología , Motilidad Espermática/efectos de los fármacos , Espermatozoides/efectos de los fármacos , Espermatozoides/fisiología
6.
Food Chem ; 320: 126652, 2020 Aug 01.
Artículo en Inglés | MEDLINE | ID: mdl-32229399

RESUMEN

Enzymatic desulfation using arylsulfatase provides an attractive approach to improve agar quality. We have previously characterized a functional arylsulfatase from Pseudoalteromonas carrageenovora. To further improve its enzymatic performance, we isolated a mutant arylsulfatase of K253Q with improved enzyme activity from a random mutant library. Compared to wild-type arylsulfatase (WT), K253Q showed 33% increase in enzyme activity, with optimal temperature and pH of 55 °C and 8.0, respectively. K253Q demonstrated better substrate binding ability with lower Km value. Structure analysis indicated that a combination of the additional hydrogen bond and the enhanced substrate binding affinity could account for the improved enzyme activity of K253Q. K253Q exhibited about 54% sulfate removal against agar, resulting in additional 8% increase in 3,6-AG content and 20% increase in gel strength compared to WT. Scanning electron microscopy showed that K253Q treatment led to a stronger crosslinking structure of agar.


Asunto(s)
Agar/química , Arilsulfatasas/genética , Arilsulfatasas/metabolismo , Pseudoalteromonas/enzimología , Evolución Molecular Dirigida , Biblioteca de Genes , Concentración de Iones de Hidrógeno , Mutación , Sulfatos/aislamiento & purificación , Sulfatos/metabolismo , Temperatura
7.
Artículo en Inglés | MEDLINE | ID: mdl-32251989

RESUMEN

Alternaria mycotoxins, such as tenuazonic acid (TeA), altenuene (ALT), alternariol (AOH), tentoxin (TEN) and alternariol monomethyl ether (AME) are frequently found in foods and may pose a potential risk to human health. Human biomonitoring can help measure our exposure to these mycotoxins, and help us determine if the exposure is changing over time. In this study, a simple liquid-liquid extraction sample preparation procedure followed by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) was developed for the simultaneous analysis of five Alternaria mycotoxins in human urine. High recoveries (92.7-103.2%) were obtained for all the tested mycotoxins with relative standard deviations (RSDs, %) of less than 6.4%. The limits of quantification (LOQs) for the analytes in urine ranged from 0.001 to 0.05 ng/mL. The method was successfully applied to investigate the levels of five Alternaria mycotoxins from 135 volunteers. In all of the samples, at least one Alternaria mycotoxin was detected. TeA, AME and AOH were the predominant Alternaria mycotoxins, and the detection rates were 85.9%, 96.3% and 51.9%, respectively.


Asunto(s)
Alternaria/química , Micotoxinas/orina , Arilsulfatasas/química , Cromatografía Líquida de Alta Presión , Contaminación de Alimentos/análisis , Glucuronidasa/química , Humanos , Lactonas/orina , Límite de Detección , Extracción Líquido-Líquido , Péptidos Cíclicos/orina , Espectrometría de Masas en Tándem , Ácido Tenuazónico/orina
8.
Clin Lab ; 66(3)2020 03 01.
Artículo en Inglés | MEDLINE | ID: mdl-32162878

RESUMEN

BACKGROUND: Rapid and accurate diagnosis of mucopolysaccharidoses (MPS) is still a challenge due to poor access to screening and diagnostic methods and to their extensive clinical heterogeneity. The aim of this work is to perform laboratory biochemical testing for confirming the diagnosis of mucopolysaccharidosis (MPS) for the first time in Morocco. METHODS: Over a period of twelve months, 88 patients suspected of having Mucopolysaccharidosis (MPS) were referred to our laboratory. Quantitative and qualitative urine glycosaminoglycan (GAG) analyses were performed, and enzyme activity was assayed on dried blood spots (DBS) using fluorogenic substrates. Enzyme activity was measured as normal, low, or undetectable. RESULTS: Of the 88 patients studied, 26 were confirmed to have MPS; 19 MPS I (Hurler syndrome; OMIM #607014/Hurler-Scheie syndrome; OMIM #607015), 2 MPS II (Hunter syndrome; OMIM #309900), 2 MPS IIIA (Sanfilippo syndrome; OMIM #252900), 1 MPS IIIB (Sanfilippo syndrome; OMIM #252920) and 2 MPS VI (Maroteaux-Lamy syndrome; OMIM #253200). Parental consanguinity was present in 80.76% of cases. Qualitative urinary glycosaminoglycan (uGAGs) assays showed abnormal profiles in 31 cases, and further quantitative urinary GAG evaluation and Thin Layer Chromatography (TLC) provided important additional information about the likely MPS diagnosis. The final diagnosis was confirmed by specific enzyme activity analysis in the DBS samples. CONCLUSIONS: The present study shows that the adoption of combined urinary substrate analysis and enzyme assays using dried blood spots can facilitate such diagnosis, offer an important tool for an appropriate supporting care, and a specific therapy, when available.


Asunto(s)
Mucopolisacaridosis/diagnóstico , Mucopolisacaridosis/orina , Urinálisis , Adolescente , Arilsulfatasas/metabolismo , Arilsulfatasas/orina , Niño , Preescolar , Cromatografía en Capa Delgada , Pruebas con Sangre Seca/economía , Pruebas con Sangre Seca/métodos , Femenino , Glicosaminoglicanos/análisis , Glicosaminoglicanos/metabolismo , Humanos , Iduronidasa/metabolismo , Iduronidasa/orina , Masculino , Marruecos , Mucopolisacaridosis/enzimología , Mucopolisacaridosis/metabolismo , Proyectos Piloto , Urinálisis/economía , Urinálisis/métodos
9.
Ecotoxicol Environ Saf ; 192: 110264, 2020 Apr 01.
Artículo en Inglés | MEDLINE | ID: mdl-32035397

RESUMEN

Copper (Cu) mining has to address a critical environmental issue related to the disposal of heavy metals and metalloids (HMs). Due to their deleterious effects on living organisms, Cu and arsenic (As) have gained global attention, and thus their monitoring in the environment is an important task. The aims of this study were: 1) to evaluate the alteration of soil enzyme activities (EAs) and soil microbial functional diversity with Cu/As contamination, and 2) to select the most reliable biochemical indicators of Cu/As contamination. A twelve-week soil experiment was performed with four increasing levels of Cu, As, and Cu/As from 150/15 to 1000/100 mg Cu/As kg-1. Soil enzyme activities and soil community-level physiological profile (CLPP) using MicroResp™ were measured during the experiment. Results showed reduced EAs over time with increasing Cu and Cu/As levels. The most Cu-sensitive EAs were dehydrogenase, acid phosphatase, and arylsulfatase, while arginine ammonification might be related to the resilience of soil microbial communities due to its increased activity in the last experimental times. There was no consistent response to As contamination with reduced individual EAs at specific sampling times, being urease the only EA negatively affected by As. MicroResp™ showed reduced carbon (C) substrate utilization with increasing Cu levels indicating a community shift in C acquisition. These results support the use of specific EAs to assess the environmental impact of specific HMs, being also the first assessment of EAs and the use of CLPP (MicroResp™) to study the environmental impact in Cu/As contaminated soils.


Asunto(s)
Arsénico/farmacología , Cobre/farmacología , Microbiología del Suelo , Contaminantes del Suelo/farmacología , Fosfatasa Ácida/metabolismo , Arilsulfatasas/metabolismo , Oxidorreductasas/metabolismo , Suelo/química , Ureasa/metabolismo
10.
Biosci. j. (Online) ; 36(1): 1-16, jan./feb. 2020. tab, ilus
Artículo en Inglés | LILACS | ID: biblio-1049184

RESUMEN

The objective of this study was to evaluate the influence of agroforestry systems of different ages (AFS1: one-year old; AFS5: five-years old) on the biological attributes of soil; the following systems were used for comparison: a slash-and-burn (SBF) farming area, Caatinga which has been undergoing regeneration for 6 years (CaR6), and native Caatinga (NCa) in Brazil. Enzyme activity, abundance and composition of arbuscular mycorrhizal fungi (AMF), and production of glomalin-related soil proteins (GRSP) were evaluated at soil depths of 0­0.05 m. AMF species composition in the AFS was more similar to that in the NCa than in the SBF and CaR6 systems. In the rainy season, sporulation was most abundant in the AFS-1, CaR6, and SBF systems, whereas GRSP concentrations were highest in the AFS5 during the dry season. Acid phosphatase and arylsulfatase enzyme activity was lower in the AFS1 soils than in the NCa and SBF soils (rainy period), and levels of ß-glucosidase and fluorescein diacetate hydrolysis in the AFS were equal to or higher than those in the NCa in the dry season but lower in the rainy season. AFS thus appear to promote the maintenance of soil biological quality, and may be more sustainable than SBF farming systems in the Brazilian Caatinga over the long term.


O objetivo do estudo foi avaliar a influência de sistemas agroflorestais (AFS1: um ano de idade; AFS5: cinco anos de idade), nos atributos biológicos do solo usando como referência, uma área de agricultura de corte e queima (SBF), Caatinga em regeneração há 6 anos (CaR6), e Caatinga nativa (NCa), in Brasil. A atividade enzimática, a abundância e composição dos fungos micorrízicos arbusculares (AMF), e a produção de proteína do solo relacionada à glomalina (GRSP) foram avaliados, na profundidade de 0-5 cm do solo. A composição das espécies de AMF nos AFS foi mais semelhante a observada na NCa, do que os sistemas SBF e CaR6. Na estação chuvosa, a esporulação foi mais abundante em AFS-1, CaR6 and SBF quando comparada as outras áreas, enquanto a GRSP apresentou maiores teores no AFS5 no período seco. AFS1 apresentou atividade da fosfatase ácida e arilsulfatase inferiores tanto a NCa quanto a SBF, no período chuvoso. No período seco, a atividade de ß-glicosidase e a hidrólise do diacetato de fluoresceína (FDA) na AFS foram iguais ou superiores a Nca, mas menor no período chuvoso. Verifica-se que os AFS são potenciais para a manutenção da qualidade biológica do solo, podendo, em longo prazo, serem mais sustentáveis que a SBF, em ambiente de Caatinga.


Asunto(s)
Arilsulfatasas , Suelo , Fosfatasa Ácida , Glicósido Hidrolasas
11.
Anal Biochem ; 588: 113474, 2020 01 01.
Artículo en Inglés | MEDLINE | ID: mdl-31614116

RESUMEN

With Escherichia coli alkaline phosphatase (ECAP) as the tag fused to the N-terminus of Pseudomonas Aeruginosa arylsulfatase (PAAS) and its mutants via a flexible linker, the comparison of the activity ratios of an applicable enzyme and its mutants to a suitable enzyme tag in cell lysates of their fused forms was tested for high-throughput (HTP) screening of mutants. After both the induced expression of a fused form and alkaline lysis of the transformed cells in microplate wells, HTP assay of the activities of ECAP and PAAS/mutant was realized via spectrophotometric-dual-enzyme-simultaneous-assay to derive their activity ratio. The successful induced expression of fused forms required ECAP activities higher than 5.3 U/L in cell lysates. Of three representative fused PAAS/mutants in cell lysates, there were similar proteolytic fragments and the comparison of their activity ratios greatly enhanced the recognition of weakly positive mutants. After saturation mutagenesis at M72 of the fused PAAS, the activity ratios of PAAS/mutants to ECAP in cell lysates of their fused forms were proportional to specific activities of their non-fused counterparts in cell lysates by an immunoturbidimetric assay. Therefore, the proposed strategy was absorbing for both HTP screening of mutants and HTP elucidation of sequence-activity relationship of applicable enzymes.


Asunto(s)
Fosfatasa Alcalina , Arilsulfatasas/química , Pruebas de Enzimas/métodos , Ensayos Analíticos de Alto Rendimiento/métodos , Proteínas Recombinantes de Fusión/química , Fosfatasa Alcalina/química , Fosfatasa Alcalina/genética , Escherichia coli/enzimología , Mutación , Pseudomonas aeruginosa/enzimología
12.
Sci Total Environ ; 703: 135536, 2020 Feb 10.
Artículo en Inglés | MEDLINE | ID: mdl-31759726

RESUMEN

Arylsulfatase and ß-glucuronidase are two important enzymes that are responsible for deconjugation of estrogen conjugates. It is important to keep estrogen conjugates intact during sample collection and storage, while the effective inhibition conditions for arylsulfatase and ß-glucuronidase remain unknown. To elucidate these conditions, inhibition experiments were performed by adding several inhibitors or by introducing extreme pH conditions. This work confirms that arylsulfatase and ß-glucuronidase can tolerate some extremes, including high concentrations of mercury dichloride, ethanol, and EDTA, while low pH (<3) or high pH (>11) can effectively inhibit their activities. The high tolerance of arylsulfatase and ß-glucuronidase for mercury dichloride explains why estrogen conjugates in wastewater samples were deconjugated, even in the extremely unfavorable condition with a high concentration of mercury dichloride. Although low pH (<3) can effectively inhibit arylsulfatase/ß-glucuronidase, deconjugation of sulfate conjugates by acid hydrolysis readily occurs; thus, a high pH of 11 is an appropriate storage condition for the effective inhibition of arylsulfatase/ß-glucuronidase. This appropriate storage condition was confirmed and validated with diluted and sterilized activated sludge samples in which arylsulfatase/ß-glucuronidase inhibition was effective for 48 h at room temperature and with a high pH of 11. The developed appropriate storage condition for effective inhibition of arylsulfatase/ß-glucuronidase has wide application potential not only for estrogen conjugates but also for all conjugates of other organic micropollutants.


Asunto(s)
Arilsulfatasas , Monitoreo del Ambiente/métodos , Glucuronidasa , Glucurónidos , Sulfatos , Aguas Residuales/química , Inhibidores Enzimáticos
13.
Front Biosci (Landmark Ed) ; 25: 760-780, 2020 01 01.
Artículo en Inglés | MEDLINE | ID: mdl-31585916

RESUMEN

Proteoglycans are essential constituents of tissue- and organ microenvironments, modulating both the structural scaffolds that surround cells as well as signaling cues that determine cellular phenotype. An important modification of proteoglycans is the sulfation of the monosaccharides that comprise them. Sulfates are added by sulfotransferases and desulfation occurs through the action of sulfatases. In this essay, we examine the biochemistry of a conserved family of desulfating enzymes known as arylsulfatases. A subset of these enzymes mediates the desulfation of proteoglycans. We review the consequences of their aberrant expression in the light of carcinogenesis and carcinomatosis: the dissemination of cancer cells. A closer understanding of their cellular-molecular roles reveals their promise for future strategies for cancer therapy.


Asunto(s)
Carcinogénesis , Proteoglicanos/metabolismo , Sulfatasas/metabolismo , Sulfatos/metabolismo , Sulfotransferasas/metabolismo , Animales , Arilsulfatasas/metabolismo , Transición Epitelial-Mesenquimal , Humanos , Transducción de Señal
14.
Environ Monit Assess ; 192(1): 20, 2019 Dec 09.
Artículo en Inglés | MEDLINE | ID: mdl-31820108

RESUMEN

The concept of the study resulted from the lack of accurate data on the toxicity of bisphenol F (BPF) coinciding with the need for immediate changes in the global economic policy eliminating the effects of environmental contamination with bisphenol A (BPA). The aim of the experiment was to determine the scale of the previously unstudied inhibitory effect of BPF on soil biochemical activity. To this end, in a soil subjected to increasing BPF pressure at three contamination levels of 0, 5, 50 and 500 mg BPF kg-1 DM, responses of soil enzymes, dehydrogenases, catalase, urease, acid phosphatase, alkaline phosphatase, arylsulphatase and ß-glucosidase, were examined. Moreover, the study suggested a potentially effective way of biostimulating the soil by means of bioaugmentation with a consortium of four bacterial species: Pseudomonas umsongensis, Bacillus mycoides, Bacillus weihenstephanensis and Bacillus subtilis, and the following fungal species: Mucor circinelloides, Penicillium daleae, Penicillium chrysogenum and Aspergillus niger. It was found that BPF was a controversial BPA analogue due to the fact that it contributed to the inhibition of all the enzyme activities. Dehydrogenases proved to be the most sensitive to bisphenol contamination of the soil. The addition of 5 mg BPF kg-1 DM of soil triggered an escalation of the inhibition comparable to that for the other enzymes only after exposing them to the effects of 50 and 500 mg BPF kg-1 DM of soil. Moreover, BPF generated low activity of urease, acid phosphatase, alkaline phosphatase and ß-glucosidase. Bacterial inoculum increased the activity of urease, ß-glucosidase, catalase and alkaline phosphatase. Seventy-six percent of BPF underwent biodegradation during the 5 days of the study.


Asunto(s)
Compuestos de Bencidrilo/análisis , Biodegradación Ambiental , Monitoreo del Ambiente , Fenoles/análisis , Microbiología del Suelo , Contaminantes del Suelo/análisis , Arilsulfatasas , Bacillus/efectos de los fármacos , Bacterias/efectos de los fármacos , Hongos/efectos de los fármacos , Oxidorreductasas , Suelo , Estrobilurinas , Ureasa
15.
Environ Sci Pollut Res Int ; 26(36): 37054-37069, 2019 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-31745783

RESUMEN

This study analysed the scale of bisphenol S (BPS) toxicity to the soil biochemical activity and is part of a wider effort to find solutions to restore the global soil environment balance, including elimination of the effects of ecosystem pollution with BPA, of which BPS is a significant analogue. However, since there has been no research on the effect of BPS on soil health, the objective of the study was pursued based on increasing the levels of soil contamination with the bisphenol 0, 5, 50 and 500 mg BPS kg-1 DM of soil and by observing the response of seven soil enzymes: dehydrogenases, catalase, urease, acid phosphatase, alkaline phosphatase, arylsulphatase and ß-glucosidase to the growing BPS pressure. The potential negative effect of bisphenol S was offset by bioaugmentation with a bacteria consortium-Pseudomonas umsongensis, Bacillus mycoides, Bacillus weihenstephanensis and Bacillus subtilis-and a fungi consortium Mucor circinelloides, Penicillium daleae, Penicillium chrysogenum and Aspergillus niger. BPS was found to be a significant inhibitor of the soil enzymatic activity and, in consequence, its fertility. Dehydrogenases and acid phosphatase proved to be the most susceptible to BPS pressure. Bioaugmentation with a bacteria consortium offset the negative effect of 500 mg BPS kg-1 DM of soil by inducing an increase in the activity of acid phosphatase and alkaline phosphatase, whereas the fungi consortium stimulated the activity of ß-glucosidase and acid phosphatase. A spectacular dimension of bisphenol S inhibition corresponded with both the spring rape above-ground parts and root development disorders and the content of Ca and K in them. The BPS level in soil on day 5 of the experiment decreased by 61% and by another 19% on day 60.


Asunto(s)
Biodegradación Ambiental , Consorcios Microbianos , Fenoles/análisis , Microbiología del Suelo , Contaminantes del Suelo/análisis , Sulfonas/análisis , Arilsulfatasas , Bacterias/efectos de los fármacos , Compuestos de Bencidrilo , Ecosistema , Contaminación Ambiental/análisis , Hongos/efectos de los fármacos , Oxidorreductasas , Fenoles/toxicidad , Suelo/química , Contaminantes del Suelo/toxicidad , Sulfonas/toxicidad , Ureasa/análisis
16.
Environ Pollut ; 252(Pt B): 1429-1438, 2019 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-31265953

RESUMEN

The aim of the work was to determine the trend, intensity and changes of selected microbial and phytotoxic parameters of degraded soil in the area of former sulphur mine reclaimed by post-flotation lime (PFL), sewage sludge (SS), mineral wool (MW- mixed with soil, MWP-pad) and mineral fertilizer (NPK). The following parameters: number of proteolytic bacteria and fungi, ammonification, nitrification, activities of alkaline phosphatase and arylsulphatase Lepidium sativum growth index (GI) and phenolic compounds were analysed in the soil in second and third year of the experiment. The addition of the SS separately or in combination with other remediation agents was found to be the most valuable for the number of microorganisms, intensification of nitrification process and enzymatic activities. In objects where other materials were added without sewage sludge, the inhibition of fungal growth as well as alkaline phosphatase and arylsulphatase activities was observed, however the inhibitory effect declined with time. The observed increase of GI shows the long-term, positive effect of treatments on soil properties concerning plant growth. The use of lime and lime together with sewage sludge contributed to the decrease in the content of phenolic compounds in the reclaimed soil.


Asunto(s)
Bacterias/crecimiento & desarrollo , Biodegradación Ambiental , Hongos/crecimiento & desarrollo , Lepidium sativum/crecimiento & desarrollo , Aguas del Alcantarillado/química , Contaminantes del Suelo/análisis , Azufre/análisis , Fosfatasa Alcalina/metabolismo , Arilsulfatasas/metabolismo , Fertilizantes/análisis , Nitrificación/fisiología , Fenol/análisis , Suelo/química , Microbiología del Suelo
17.
Microscopy (Oxf) ; 68(3): 243-253, 2019 Jun 01.
Artículo en Inglés | MEDLINE | ID: mdl-30860257

RESUMEN

This study was designed to observe osteoclasts in the rat femora by light and electron microscopic cytochemistry for nicotinamide adenine dinucleotide phosphatase (NADPase) and arylsulfatase, and scanning electron microscopy using osmium maceration to assess the three-dimensional morphology of the Golgi apparatus in osteoclasts. The Golgi apparatus showed strong NADPase activity and surrounded each nucleus with the cis-side facing the nucleus. The Golgi apparatus could be often traced for a length of 20 µm or longer. Observations of serial semi-thin sections confirmed that a single line of reaction products (=lead precipitates) intervened somewhere between any two neighboring nuclei. The nuclear membrane showed strong arylsulfatase activity as well as rough endoplasmic reticulum and lysosomes. Scanning electron microscopy showed that the Golgi apparatus covered the nucleus in a porous sheet-like configuration. Under magnification, the cis-most saccule showed a sieve-like configuration with fine fenestrations. The saccules decreased fenestration numbers toward the trans-side and displayed a more plate-like appearance. The above findings indicate the following. (1) The Golgi saccules of osteoclasts have a three-dimensional structure comparable with that generally seen in other cell types. (2) The Golgi apparatus forms a porous multi-spherical structure around nuclei. Within the structure, in most cases a Golgi stack partitions the room into several compartments in each of which a nucleus fits. (3) The nuclear membrane synthesizes some kinds of proteins more stably and sufficiently than the rough endoplasmic reticulum. Consequently, the Golgi apparatus accumulates around nuclei with the cis-side facing the nucleus.


Asunto(s)
Arilsulfatasas/metabolismo , Aparato de Golgi/ultraestructura , NAD/química , Osteoclastos/ultraestructura , Pirofosfatasas/metabolismo , Animales , Retículo Endoplásmico Rugoso/metabolismo , Aparato de Golgi/metabolismo , Lisosomas/metabolismo , Masculino , Microscopía Electrónica de Rastreo , Membrana Nuclear/metabolismo , Osmio/química , Ratas , Ratas Wistar
18.
Biochemistry ; 58(10): 1363-1378, 2019 03 12.
Artículo en Inglés | MEDLINE | ID: mdl-30810299

RESUMEN

Pseudomonas aeruginosa arylsulfatase (PAS) hydrolyzes sulfate and, promiscuously, phosphate monoesters. Enzyme-catalyzed sulfate transfer is crucial to a wide variety of biological processes, but detailed studies of the mechanistic contributions to its catalysis are lacking. We present linear free energy relationships (LFERs) and kinetic isotope effects (KIEs) of PAS and analyses of active site mutants that suggest a key role for leaving group (LG) stabilization. In LFERs PASWT has a much less negative Brønsted coefficient (ßleaving groupobs-Enz = -0.33) than the uncatalyzed reaction (ßleaving groupobs = -1.81). This situation is diminished when cationic active site groups are exchanged for alanine. The considerable degree of bond breaking during the transition state (TS) is evidenced by an 18Obridge KIE of 1.0088. LFER and KIE data for several active site mutants point to leaving group stabilization by active site K375, in cooperation with H211. 15N KIEs and the increased sensitivity to leaving group ability of the sulfatase activity in neat D2O (Δßleaving groupH-D = +0.06) suggest that the mechanism for S-Obridge bond fission shifts, with decreasing leaving group ability, from charge compensation via Lewis acid interactions toward direct proton donation. 18Ononbridge KIEs indicate that the TS for PAS-catalyzed sulfate monoester hydrolysis has a significantly more associative character compared to the uncatalyzed reaction, while PAS-catalyzed phosphate monoester hydrolysis does not show this shift. This difference in enzyme-catalyzed TSs appears to be the major factor favoring specificity toward sulfate over phosphate esters by this promiscuous hydrolase, since other features are either too similar (uncatalyzed TS) or inherently favor phosphate (charge).


Asunto(s)
Arilsulfatasas/metabolismo , Fosfatos/química , Sulfatos/química , Arilsulfatasas/genética , Catálisis , Dominio Catalítico , Hidrólisis , Cinética , Organofosfatos/química , Compuestos Organofosforados/química , Fosfatos/metabolismo , Pseudomonas aeruginosa/metabolismo , Especificidad por Sustrato/genética , Especificidad por Sustrato/fisiología , Sulfatasas/química , Sulfatos/metabolismo
19.
Bioorg Med Chem ; 27(6): 955-962, 2019 03 15.
Artículo en Inglés | MEDLINE | ID: mdl-30738652

RESUMEN

Sulfatases hydrolyze sulfated metabolites to their corresponding alcohols and are present in all domains of life. These enzymes have found major application in metabolic investigation of drugs, doping control analysis and recently in metabolomics. Interest in sulfatases has increased due to a link between metabolic processes involving sulfated metabolites and pathophysiological conditions in humans. Herein, we present the first comprehensive substrate specificity and kinetic analysis of the most commonly used arylsulfatase extracted from the snail Helix pomatia. In the past, this enzyme has been used in the form of a crude mixture of enzymes, however, recently we have purified this sulfatase for a new application in metabolomics-driven discovery of sulfated metabolites. To evaluate the substrate specificity of this promiscuous sulfatase, we have synthesized a series of new sulfated metabolites of diverse structure and employed a mass spectrometric assay for kinetic substrate hydrolysis evaluation. Our analysis of the purified enzyme revealed that the sulfatase has a strong preference for metabolites with a bi- or tricyclic aromatic scaffold and to a lesser extent for monocyclic aromatic phenols. This metabolite library and mass spectrometric method can be applied for the characterization of other sulfatases from humans and gut microbiota to investigate their involvement in disease development.


Asunto(s)
Arilsulfatasas/metabolismo , Caracoles Helix/enzimología , Animales , Caracoles Helix/metabolismo , Hidrólisis , Cinética , Espectrometría de Masas , Especificidad por Sustrato
20.
J Mol Neurosci ; 67(3): 472-476, 2019 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-30656493

RESUMEN

Blepharospasm (BSP) is a sub-phenotype of focal dystonia. A few genetic risk factors are considered to be implicated in the risk of developing BSP. There is recent evidence, based on results from GWAS and meta-analyses, to suggest that arylsulfatase G (ARSG), and more specifically rs11655081, is implicated in focal dystonia. The aim of the present study was to evaluate the effect of rs11655081 ARSG on BSP. A Greek cohort, which consisted of 206 BSP patients and an equal number of healthy controls, was genotyped for rs11655081. Only a marginal trend for the association between rs11655081 and the risk of BSP was found in the over-dominant model of inheritance [odds ratio, OR (95% confidence interval, CI): 0.64 (0.38-1.07), p = 0.088]. It is rather unlikely that rs11655081 across ARSG is a major genetic risk contributor for BSP.


Asunto(s)
Arilsulfatasas/genética , Blefaroespasmo/genética , Polimorfismo de Nucleótido Simple , Humanos
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