RESUMEN
The small nuclear RNA (snRNA) activating protein complex (SNAPc) is essential for transcription of genes that encode the snRNAs. Drosophila melanogaster SNAPc (DmSNAPc) consists of three subunits (DmSNAP190, DmSNAP50 and DmSNAP43) that form a stable complex that recognizes an snRNA gene promoter element called the PSEA. Although all three subunits are required for sequence-specific DNA binding activity, only DmSNAP190 possesses a canonical DNA binding domain consisting of 4.5 tandem Myb repeats homologous to the Myb repeats in the DNA binding domain of the Myb oncoprotein. In this study, we use site-specific protein-DNA photo-cross-linking technology followed by site-specific protein cleavage to map domains of DmSNAP190 that interact with specific phosphate positions in the U6 PSEA. The results indicate that at least two DmSNAP190 Myb repeats contact the DNA in a significantly different manner when DmSNAPc binds to a U6 PSEA versus a U1 PSEA, even though the two PSEA sequences differ at only 5 of 21 nucleotide positions. The results are consistent with a model in which the specific DNA sequences of the U1 and U6 PSEAs differentially alter the conformation of DmSNAPc, leading to the subsequent recruitment of different RNA polymerases to the U1 and U6 gene promoters.
Asunto(s)
Proteínas de Unión al ADN/química , Proteínas de Unión al ADN/metabolismo , Proteínas de Drosophila/química , Proteínas de Drosophila/metabolismo , Regiones Promotoras Genéticas , ARN Nuclear Pequeño/genética , Secuencia de Bases , ADN/química , Modelos Genéticos , Unión Proteica , Estructura Terciaria de ProteínaRESUMEN
A microbiological algorithm has been developed to analyze beach water samples for the determination of viable colony forming units (CFU) of Staphylococcus aureus (S. aureus). Membrane filtration enumeration of S. aureus from recreational beach waters using the chromogenic media CHROMagar SA alone yields a positive predictive value (PPV) of 70%. Presumptive CHROMagar SA colonies were confirmed as S. aureus by 24-hour tube coagulase test. Combined, these two tests yield a PPV of 100%. This algorithm enables accurate quantitation of S. aureus in seawater in 72 hours and could support risk-prediction processes for recreational waters. A more rapid protocol, utilizing a 4-hour tube coagulase confirmatory test, enables a 48-hour turnaround time with a modest false negative rate of less than 10%.
