Three dimensional structure of the MqsR:MqsA complex: a novel TA pair comprised of a toxin homologous to RelE and an antitoxin with unique properties.

One mechanism by which bacteria survive environmental stress is through the formation of bacterial persisters, a sub-population of genetically identical quiescent cells that exhibit multidrug tolerance and are highly enriched in bacterial toxins. Recently, the Escherichia coli gene mqsR (b3022) was identified as the gene most highly upregulated in persisters. Here, we report multiple individual and complex three-dimensional structures of MqsR and its antitoxin MqsA (B3021), which reveal that MqsR:MqsA form a novel toxin:antitoxin (TA) pair. MqsR adopts an alpha/beta fold that is homologous with the RelE/YoeB family of bacterial ribonuclease toxins. MqsA is an elongated dimer that neutralizes MqsR toxicity. As expected for a TA pair, MqsA binds its own promoter. Unexpectedly, it also binds the promoters of genes important for E. coli physiology (e.g., mcbR, spy). Unlike canonical antitoxins, MqsA is also structured throughout its entire sequence, binds zinc and coordinates DNA via its C- and not N-terminal domain. These studies reveal that TA systems, especially the antitoxins, are significantly more diverse than previously recognized and provide new insights into the role of toxins in maintaining the persister state.

Functional characterization of Vip3Ab1 and Vip3Bc1: Two novel insecticidal proteins with differential activity against lepidopteran pests.

In this work, we characterized 2 novel insecticidal proteins; Vip3Ab1 and Vip3Bc1. These proteins display unique insecticidal spectra and have differential rates of processing by lepidopteran digestive enzymes. Furthermore, we have found that both proteins exist as tetramers in their native state before and after proteolysis. In addition, we expressed truncated forms and protein chimeras to gain a deeper understanding of toxin specificity and stability. Our study confirms a role for the C-terminal 65 kDa domain in directing insect specificity. Importantly, these data also indicate a specific interaction between the 20 kDa amino terminus and 65 kDa carboxy terminus, after proteolytic processing. We demonstrate the C-terminal 65 kDa to be labile in native proteolytic conditions in absence of the 20 kDa N-terminus. Thus, the 20 kDa fragment functions to provide stability to the C-terminal domain, which is necessary for lethal toxicity against lepidopteran insects.