Conserved lysine residues in decorin binding proteins of Borrelia garinii are critical in adhesion to human brain microvascular endothelial cells.

Lyme borreliosis is a tick-borne disease caused by Borrelia burgdorferi sensu lato spirochetes (Lyme borreliae). When the disease affects the central nervous system, it is referred to as neuroborreliosis. In Europe, neuroborreliosis is most often caused by Borrelia garinii. Although it is known that in the host Lyme borreliae spread from the tick bite site to distant tissues via the blood vasculature, the adherence of Lyme borreliae to human brain microvascular endothelial cells has not been studied before. Decorin binding proteins are adhesins expressed on Lyme borreliae. They mediate the adhesion of Lyme borreliae to decorin and biglycan, and the lysine residues located in the binding site of decorin binding proteins are important to the binding activity. In this study, we show that lysine residues located in the canonical binding site can also be found in decorin binding proteins of Borrelia garinii, and that these lysines contribute to biglycan and decorin binding. Most importantly, we show that the lysine residues are crucial for the binding of Lyme borreliae to decorin and biglycan expressing human brain microvascular endothelial cells, which in turn suggests that they are involved in the pathogenesis of neuroborreliosis.

Immobilization of bromelain on cobalt-iron magnetic nanoparticles (CoFe2O4) for casein hydrolysis / Inmovilización de bromelina sobre nanopartículas magnéticas de cobalto-hierro (CoFe2O4) para la hidrólisis de caseína / Inmovilización de bromelina sobre nanopartículas magnéticas de cobalto-hierro (CoFe2O4) para a hidrólise de caseína

Abstract By means of recycling an enzyme, bromelain was used in casein hydrolysis facilitated by a nanobiocatalyst consisting of bromelain, CoFe2O4 magnetic nanoparticles, chitosan, and glutaraldehyde. Bromelain was immobilized on the chitosan cobalt-magnetite nanoparticle surface via covalent bonds to form the nanobiocatalyst. Immobilized bromelain showed 77% immobilization binding, retaining 85 ± 2% of the initial catalytic activity. Nanoparticles and immobilized bromelain were characterized using UV-Vis and IR spectroscopies, X-ray, square wave voltammetry (SWV), cyclic voltammetry (CV), vibrating-sample magnetization (VSM), and transmission electron microscope (TEM). The Michaelis-Menten constant (KM) and VMAX of the free and immobilized enzyme were calculated: KM = 2.1 ± 0.18 mM and 1.8 mM, respectively and VMAX = 6.08 x 10-2 ± 2.1 x 10-2 U/min and 6.46 ± 0.91 U/min, respectively. The thermal stability of the free enzyme was higher than the immobilized enzyme: 95-98% and 83-87%, respectively. An optimum pH of 6 and a temperature of 20 °C were determined in both cases. Immobilized bromelain maintained 50% of the initial catalytic activity after the fifth use. The immobilized bromelain proved to be effective and reusable for casein hydrolysis. As novel contribution the characterization by VOC and CV was carried out.

Resumen Se investigó la reutilización de la bromelina inmovilizada sobre nanopartículas magnéticas de CoFe2O4 para la hidrólisis de caseína. La inmovilización se efectuó covalentemente en un catalizador de nanopartículas de hierro y cobalto recubiertas con quitosano, glutaraldehído y bromelina. La bromelina alcanzó un máximo de inmovilización del 77%, conservando el 85 ± 2% de su actividad catalítica inicial a los 30 min. La caracterización del catalizador se realizó por espectrofotometría IR y UV-Vis, microscopía electrónica de transmisión (TEM), difracción de rayos X, voltametría de onda cuadrada (VOC), voltametría cíclica (VC) y magnetización de muestra vibrante (VSM). Los parámetros cinéticos KM y VMAX de la enzima libre e inmovilizada fueron: KM = 2,1 ± 0,18 mM y 1,8 ± 0,16 mM, respectivamente y VMAX = 6,08 x 10-2 ± 2,1 x 10-2 U/min y 6,46 ± 0,91 U/min, respectivamente. La estabilidad térmica de la enzima libre fue mayor que la de enzima inmovilizada: 95-98% y 83-87%, respectivamente. Se determinó un pH óptimo de 6 y temperatura de 20 °C en ambos casos. La bromelina inmovilizada mantuvo el 50% de su actividad catalítica hasta el quinto uso. Como aporte novedoso se realizó, en este estudio se realizó la caracterización por VOC y VC.

Resumo Foi estudado o reuso da enzima bromelina imobilizada sobre nano partículas magnéticas de CoFe2O4, para a hidrólise de caseína. A imobilização efetuou-se covalentemente em um catalisador de nano partículas de cobalto e ferro recobertas com quitosano, glutaraldehido e bromelina. A bromelina atingiu seu máximo de imobilização do 77% aos 30 minutos, conservando o 85 ± 2% da sua atividade catalítica inicial. A caraterização foi feita mediante espectrofotometria IV e UV-Vis, microscopia eletrônica de transmissão (TEM), difração de raios X, voltametria de onda quadrada (VOQ), voltametria cíclica (VC) e por magnetização de amostra vibrante (VSM). Os parâmetros cinéticos KM e VMAX da enzima livre e imobilizada foram KM = 2,1 ± 0,18 mM e 1,8 8 ± 0,16mM, respeitivamente; VMAX = 6,08 x 10-2 ± 2,1 x 10-2 U/min e 6,46 ± 0,91 U/min, respectivamente. A estabilidade térmica da enzima livre (95-98%) foi maior do que a estabilidade da enzima imobilizada (83-87%), determinou-se um pH óptimo de 6 e temperatura de 20 °C em ambos os casos. A bromelina imobilizada manteve o 50% de sua atividade catalítica até o quinto uso. Como aporte inovador neste estudo apresenta-se a caraterização por VOQ e VC.

Structural and Biomolecular Analyses of Borrelia burgdorferi BmpD Reveal a Substrate-Binding Protein of an ABC-Type Nucleoside Transporter Family.

Borrelia burgdorferisensu lato, the causative agent of tick-borne Lyme borreliosis (LB), has a limited metabolic capacity and needs to acquire nutrients, such as amino acids, fatty acids, and nucleic acids, from the host environment. Using X-ray crystallography, liquid chromatography-mass spectrometry, microscale thermophoresis, and cellular localization studies, we show that basic membrane protein D (BmpD) is a periplasmic substrate-binding protein of an ABC transporter system binding to purine nucleosides. Nucleosides are essential for bacterial survival in the host organism, and these studies suggest a key role for BmpD in the purine salvage pathway of B. burgdorferi sensu lato Because B. burgdorferisensu lato lacks the enzymes required for de novo purine synthesis, BmpD may play a vital role in ensuring access to the purines needed to sustain an infection in the host. Furthermore, we show that, although human LB patients develop anti-BmpD antibodies, immunization of mice with BmpD does not confer protection against B. burgdorferi sensu lato infection.

Borrelia burgdorferi Infection in Biglycan Knockout Mice.

BACKGROUND: Borrelia burgdorferi sensu lato spirochetes (Borrelia) causing Lyme borreliosis are able to disseminate from the initial entry site to distant organs in the host. Outer-surface adhesins are crucial in the bacterial dissemination and adhesion to various tissues. Two well-characterized Borrelia adhesins, decorin-binding proteins A and B, have been shown to bind to 2 host receptors, decorin and biglycan. However, the role of biglycan in Borrelia infection has not been characterized in vivo. METHODS: We infected biglycan knockout (KO) and wild-type (WT) C3H mice with strains representing 3 Borrelia genospecies, Borrelia burgdorferi sensu stricto, Borrelia garinii, and Borrelia afzelii. The infection was monitored by measuring joint swelling, Borrelia culture, polymerase chain reaction analysis, and serologic analysis. The host immune responses were analyzed by histological scoring of the inflammation in tissues and by cytokine profiling. RESULTS: B. burgdorferi sensu stricto and B. garinii established long-term infection in mice of both genotypes, while B. afzelii failed to disseminate in KO mice. Further, the B. burgdorferi sensu stricto-infected KO mice had persistent inflammation in the joints. CONCLUSIONS: The dissemination and tissue colonization of Borrelia and the inflammatory response of the host differ in a mouse biglycan expression- and Borrelia genospecies-dependent manner.

Predicting the ligand-binding properties of Borrelia burgdorferi s.s. Bmp proteins in light of the conserved features of related Borrelia proteins.

Bacteria of the genus Borrelia cause vector-borne infections like the most important hard tick-borne disease in the northern hemisphere, Lyme borreliosis (LB), and soft tick or louse transmitted relapsing fevers (RF), prevalent in temperate and tropical areas. Borrelia burgdorferi sensu lato (s.l.) includes several genospecies and causes LB in humans. In infected patients, Borrelia burgdorferi sensu stricto (s.s.) expresses the BmpA, BmpB, BmpC and BmpD proteins. The role of these proteins in the pathogenesis of LB remains incompletely characterized, but they are, however, closely related to Treponema pallidum PnrA (Purine nucleoside receptor A), a substrate-binding lipoprotein of the ATP-binding cassette (ABC) transporter family preferentially binding purine nucleosides. Based on 3D homology modeling, the Bmp proteins share the typical fold of the substrate-binding protein family and the ligand-binding properties of BmpA, BmpB and BmpD are highly similar, whereas those of BmpC differ markedly. Nevertheless, these residues are highly conserved within the genus Borrelia and the inferred phylogenetic tree also reveals that the RF Borrelia lack BmpB proteins but has an additional Bmp protein (BmpA2) missing in LB-causing Borrelia burgdorferi s.l. Our results indicate that the Bmp proteins could bind nucleosides, although BmpC might have a different ligand-binding specificity and, therefore, a distinct function. Furthermore, the work provides a means for classifying the Bmp proteins and supports further elucidation of the roles of these proteins.

Nanocristales para degradación de un colorante contaminante / Nanocrystals for the degradation of a pollutant dye / Nanocristais para a degradação de um corante poluente

La fotoestabilidad de nanocristales (NCs) del grupo E-VI, como CdSe se aprovecha en procesos de fotocatálisis. Sin embargo, el contenido de metales tóxicos limita su aplicación en el ambiente en la remediación de residuos de las industrias textiles, de pieles y de papel. Se comparó la acción catalítica de CdS y ZnS, estabilizadas con dodecil sulfato de sodio y ácido etilendiaminotetraacético, respectivamente, para el tratamiento fotocatalítico del colorante azul de metileno (AM) con radiación UV. Las dos clases de NCs presentaron bandas de absorción desplazadas hacia 500 nm y bandas de emisión fluorescente a 430-440 nm. Estas características ópticas se atribuyen al tamaño (20-50 y 100-150 nm), confirmado por microscopía electrónica. La degradación del AM por NCs alcanzó rendimientos del 92% y 77% para ZnS y CdS respectivamente, mediante la producción fotocatalítica de radicales hidroxilos capaces de participar en procesos redox. En conclusión, NCs se aprecian como catalizadores eficientes para la remediación de AM, un colorante aromático heterocíclico de amplio uso industrial.

The photo-stability of nanocrystals (NCs) in II-VI group such as CdSe is used for photo-catalytic processes. The presence of toxic metals limits their implementation in environmental applications such as synthetic dyes treatment, which are found in textile, leather, and paper industries. NCs ZnS and CdS, stabilized in the presence of ethylendiaminetetraacetic acid and sodium dodecyl sulfate, respectively, were compared in the photo-catalytic treatment of methylene blue (MB), under UV light. The two kinds of NCs showed absorption bands displaced towards 500 nm and fluorescent emission bands around 430 nm. These optical properties are attributed to nano size as observed by scanning electron microscopy (20-50 and 100-150 nm). MB degradation by NCs reached yields up to 92% and 77% for ZnS and CdS, respectively, due to the photo-catalytic production of hydroxyl radicals capable of participating in redox processes. In conclusion, NCs catalyzed the remediation of MB, a heterocyclic aromatic dye of broad industrial use.

A fotoestabilidade do nanomateriais (NCs) do grupo II-VI como CdSe é usada em processos de fotocatálise. Mas a presença de metais tóxicos limita sua implementação no ambiente para remediar os residuos das industrias têxteis, de couro e de papel. Foi comparada a ação catalítica de ZnS e CdS, estabilizados na presenca de dodecil sulfato de sódio e ácido etilenodiamino tetra-acético, respectivamente, para o tratamento fotocatalítico de azul de metileno (AM) com radiação UV. Os dois tipos de NCs mostraram bandas de absorção perto de 500 nm e bandas de emissao fluorescente a 430-440 nm Estas propriedades ópticas são atribuídas ao tamanho (20-50 y 100-150 nm) confirmado por microscopia electrónica A degradação do AM por NCs alcancou rendimentos de 92% e 77% para ZnS e CdS respectivamente, através da produção fotocatalítica de radicais hidroxilo capazes de participar em processos redox Em conclusão NCs são vistos como catalisadores eficientes para a remediação de AM, um corante de amplia utilização industrial.