RESUMO
Operons encoding stable toxins and their labile antidote are widespread in prokaryotes and play important roles in plasmid partitioning and cellular responses to stress. One such family of toxins MazF/ChpAK/PemK encodes an endoribonuclease that inactivates cellular mRNAs by cleaving them at specific, but frequently occurring sites. Here we show that the Bacillus subtilis ydcE gene encodes a member of this family of RNases, which we have called EndoA. Overexpression of EndoA is toxic for bacterial cell growth and this toxicity is reversed by coexpression of the gene immediately upstream, ydcD. Furthermore, YdcD inhibits EndoA activity directly in vitro. EndoA has similar cleavage specificity to MazF and PemK and yields cleavage products with 3'-phosphate and 5'-hydroxyl groups, typical of EDTA-resistant degradative RNases. This is the first example of an antitoxin-toxin system in B. subtilis.
Assuntos
Bacillus subtilis/enzimologia , Endorribonucleases/antagonistas & inibidores , Endorribonucleases/genética , Óperon , Bacillus subtilis/genética , Proteínas de Bactérias/genética , Proteínas de Bactérias/fisiologia , Toxinas Bacterianas/antagonistas & inibidores , Toxinas Bacterianas/genética , Toxinas Bacterianas/metabolismo , Endorribonucleases/química , Endorribonucleases/metabolismo , Modelos Moleculares , RNA Mensageiro/metabolismo , Especificidade por SubstratoRESUMO
Crystal structures are reported for free and coenzyme A (CoA) bound forms of the YfdW protein from Escherichia coli, a representative type III CoA transferase. The structures reveal a two-domain protomer with interdomain connections forming a ring-like structure with a large central hole. Two protomers associate to form a highly intertwined dimer in which the hole of each ring is filled by the partner molecule. Each protomer binds a single CoA molecule and these CoA-binding sites are distant from one another in the dimer.
Assuntos
Coenzima A-Transferases/química , Coenzima A-Transferases/metabolismo , Coenzima A/metabolismo , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/metabolismo , Escherichia coli/enzimologia , Sítios de Ligação , Cristalografia por Raios X , Dimerização , Ligação Proteica , Estrutura Quaternária de Proteína , Estrutura Terciária de ProteínaRESUMO
Glutathione synthase catalyzes the final ATP-dependent step in glutathione biosynthesis, the formation of glutathione from gamma-glutamylcysteine and glycine. We have determined structures of yeast glutathione synthase in two forms: unbound (2.3 A resolution) and bound to its substrate gamma-glutamylcysteine, the ATP analog AMP-PNP, and two magnesium ions (1.8 A resolution). These structures reveal that upon substrate binding, large domain motions convert the enzyme from an open unliganded form to a closed conformation in which protein domains completely surround the substrate in the active site.
