RESUMEN
Cyclic di-GMP (c-di-GMP), a ubiquitous bacterial second messenger, has emerged as a key controller of several biological processes. Numbers of reports that deal with the mechanistic aspects of this second messenger have appeared in the literature. However, the lack of a reporter tag attached to the c-di-GMP at times limits the understanding of further details. In this study, we have chemically coupled N-methylisatoic anhydride (MANT) with c-di-GMP, giving rise to Mant-(c-di-GMP) or MANT-CDG. We have characterized the chemical and physical properties and spectral behavior of MANT-CDG. The fluorescence of MANT-CDG is sensitive to changes in the microenvironment, which helped us study its interaction with three different c-di-GMP binding proteins (a diguanylate cyclase, a phosphodiesterase, and a PilZ domain-containing protein). In addition, we have shown here that MANT-CDG can inhibit diguanylate cyclase activity; however, it is hydrolyzed by c-di-GMP specific phosphodiesterase. Taken together, our data suggest that MANT-CDG behaves like native c-di-GMP, and this study raises the possibility that MANT-CDG will be a valuable research tool for the in vitro characterization of c-di-GMP signaling factors.
Asunto(s)
GMP Cíclico/análogos & derivados , Colorantes Fluorescentes/síntesis química , Colorantes Fluorescentes/metabolismo , Secuencia de Aminoácidos , Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Técnicas de Química Sintética , GMP Cíclico/síntesis química , GMP Cíclico/química , GMP Cíclico/metabolismo , Transferencia Resonante de Energía de Fluorescencia , Colorantes Fluorescentes/química , Hidrólisis , Cinética , Datos de Secuencia Molecular , Reproducibilidad de los ResultadosRESUMEN
Electron transport and respiratory pathways are active in both latent and rapidly growing mycobacteria and remain conserved in all mycobacterial species. In mycobacteria, menaquinone is the sole electron carrier responsible for electron transport. Menaquinone biosynthesis pathway is found to be essential for the growth of mycobacteria. Structural analogs of the substrate or product of this pathway are found to be inhibitory for the growth of Mycobacterium smegmatis and M. tuberculosis. Several plumbagin [5-hydroxy-2-methyl-1, 4-naphthaquinone] derivatives have been analyzed for their inhibitory effects of which butyrate plumbagin was found to be most effective on M. smegmatis mc(2)155, whereas crotonate plumbagin showed greater activity on M. tuberculosis H37Rv. Effect on electron transport and respiration was demonstrated by butyrate plumbagin inhibiting oxygen consumption in M. smegmatis. Structural modifications of these molecules can further be improved upon to generate new molecules against mycobacteria.
Asunto(s)
Antituberculosos/química , Antituberculosos/farmacología , Mycobacterium/efectos de los fármacos , Naftoquinonas/química , Naftoquinonas/farmacología , Pruebas de Sensibilidad Microbiana , Mycobacterium/crecimiento & desarrollo , Infecciones por Mycobacterium/tratamiento farmacológico , Mycobacterium smegmatis/efectos de los fármacos , Mycobacterium smegmatis/crecimiento & desarrollo , Mycobacterium tuberculosis/efectos de los fármacos , Mycobacterium tuberculosis/crecimiento & desarrolloRESUMEN
Pythium campanulatum sp. nov. was isolated from some soil samples taken in the rhizosphere of maize (Zea mays) in north-eastern India. This species is characterized by the absence of zoospores and sporangia, antheridial branches wrapping around the oogonia leaving one to two campanulate antheridial cells after fertilization, and aplerotic oospores. The ITS region of its rDNA is comprised of 922 bases. This oomycete is closely related to Pythium orthogonon, Pythium nunn and Pythium toruloides. However, it has its own characteristic features and is completely devoid of zoospores. Taxonomic description of this new species and its comparison with related oomycetes, together with the sequence of the PCR-amplified internal transcribed region (spacers ITS1, ITS2, and the gene 5.8S) of its rDNA are given here.
Asunto(s)
Raíces de Plantas/microbiología , Pythium/clasificación , Pythium/aislamiento & purificación , Microbiología del Suelo , Zea mays/microbiología , Secuencia de Bases , ADN de Hongos/química , ADN de Hongos/aislamiento & purificación , ADN Espaciador Ribosómico/química , ADN Espaciador Ribosómico/aislamiento & purificación , India , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa , Pythium/citología , Pythium/genética , Homología de Secuencia de Ácido NucleicoRESUMEN
Pythium rhizosaccharum (F-1244) was isolated from soil samples taken in the rhizosphere of sugarcane (Saccharum officinarum) in the north-eastern India. This species is characterized by its smooth-walled, spherical sporangia and rarely formed sexual structures. When formed, the antheridial branches wrap around the oogonia and soon disappear after fertilization. The internal transcribed spacer (ITS) region of its rDNA is comprised of 904 bases. The taxonomical description of this new species and its comparison with related species are given here, together with the nucleotide sequences of the ITS1 and ITS2, and the 5.8S gene of its ribosomal nuclear DNA.