RESUMO
An anti-TRAP (AT) protein, a factor of previously unknown function, conveys the metabolic signal that the cellular transfer RNA for tryptophan (tRNATrp) is predominantly uncharged. Expression of the operon encoding AT is induced by uncharged tRNATrp. AT associates with TRAP, the trp operon attenuation protein, and inhibits its binding to its target RNA sequences. This relieves TRAP-mediated transcription termination and translation inhibition, increasing the rate of tryptophan biosynthesis. AT binds to TRAP primarily when it is in the tryptophan-activated state. The 53-residue AT polypeptide is homologous to the zinc-binding domain of DnaJ. The mechanisms regulating tryptophan biosynthesis in Bacillus subtilis differ from those used by Escherichia coli.
Assuntos
Bacillus subtilis/genética , Bacillus subtilis/metabolismo , Proteínas de Bactérias/metabolismo , Regulação Bacteriana da Expressão Gênica , Proteínas de Ligação a RNA/metabolismo , Fatores de Transcrição/metabolismo , Triptofano/biossíntese , Regiões 5' não Traduzidas/metabolismo , Motivos de Aminoácidos , Sequência de Aminoácidos , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/isolamento & purificação , Reagentes de Ligações Cruzadas , Eletroforese em Gel de Poliacrilamida , Glutaral , Dados de Sequência Molecular , Óperon , RNA Bacteriano/metabolismo , RNA de Transferência de Triptofano/metabolismo , Proteínas de Ligação a RNA/antagonistas & inibidores , Alinhamento de Sequência , Fatores de Transcrição/antagonistas & inibidores , Transcrição GênicaRESUMO
We have cloned and sequenced the nrd (nucleotide reductase) locus of Bacillus subtilis. The locus seems to be organized in an operon comprising four ORFs. The first three encode polypeptides highly similar to the product of the coding sequences characterizing the nrdEF operons of Enterobacteriaceae. The sequencing of the conditional lethal mutation ts-A13, localized in the nrdE cistron, and the lethality of insertional mutations targeted in the internal region of nrdE and nrdF, demonstrated the essential role of this locus. The fourth ORF, ymaB, part of the putative operon, which is not similar to any known protein, is also essential. The regulation of expression of the operon, monitored by lacZ transcriptional fusions, is similar to the regulation of the functionally relevant nrdAB operon of Escherichia coli. The operon was induced by thymidine starvation and its expression was directly or indirectly affected by RecA function. Genetic and functional analysis strongly indicates that in B. subtilis the class I ribonucleotide reductase encoded by this nrd operon is evolutionarily distant from the homologous class I enzyme of Enterobacteria.