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

Abstract

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 α/β 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.

The ParE2–PaaA2 toxin–antitoxin complex fromEscherichia coliO157 forms a heterodocecamer in solution and in the crystal

“…The antitoxin is a modular protein with a DNA-binding domain of variable fold linked to a (usually) intrinsically disordered toxin-neutralizing segment. This toxin-neutralizing segment binds to or even complements the fold of the toxin and inhibits its biochemical activity (Kamada et al, 2003;Kamada & Hanaoka, 2005;Garcia-Pino et al, 2008;Li et al, 2009;De Jonge et al, 2009;Brown et al, 2009). This activity depends on the family of toxin considered and may include poisoning DNA gyrase (Bernard & Couturier, 1992;Jiang et al, 2002), cleaving RNA in a ribosome-dependent or independent fashion (Zhang et al, 2003;Zhang & Inouye, 2011), direct ribosome inhibition (Liu et al, 2008), chemical modification of ribosomes (Vesper et al, 2011), modifying initiator tRNA or a number of other activities that directly alter the basic physiology of the cell (Yamamoto et al, 2009;Tan et al, 2010;Mutschler et al, 2011).…”

Structure of the Escherichia coli Antitoxin MqsA (YgiT/b3021) Bound to Its Gene Promoter Reveals Extensive Domain Rearrangements and the Specificity of Transcriptional Regulation

“…TA pairs are composed of two genes organized in an operon that encode an unstable, labile antitoxin and a stable toxin, which associate to form a stable complex (13)(14)(15)(16)(17). There are currently nine established toxin families (18), and similarities between toxins in fold and function are slowly emerging (19,20). However, there is much greater diversity in structure and toxin recognition among antitoxins.…”

“…Recently, we determined the crystal structures of MqsA alone and the MqsR⅐MqsA complex and showed that the mqsRA operon defines a novel family of TA systems in E. coli, in which MqsR is the toxin and MqsA is the antitoxin (19). The structure revealed that the MqsR toxin is a ribonuclease belonging to the RelE bacterial toxin family (19,34,35). MqsR is unique because it is the first toxin linked to biofilms and quorum sensing (33) and is the first toxin that, when deleted, decreases persister cell formation (31).…”

“…MqsR is unique because it is the first toxin linked to biofilms and quorum sensing (33) and is the first toxin that, when deleted, decreases persister cell formation (31). The antitoxin MqsA is even more unusual because it is the first antitoxin that requires a metal, zinc, for structural stability (19); it is the only E. coli antitoxin that is structured throughout its entire sequence; and it is the first antitoxin demonstrated to bind DNA via its C-terminal and not N-terminal domain (19, 27, 36 -39). Critically, in addition to binding its own promoter (19,34,35,40), MqsA and the MqsR⅐MqsA complex are also the only known antitoxin/TA pair that bind not only their own promoter but also the promoters of other genes that play important roles in E. coli physiology, including mcbR and spy (19) as well as cspD (31).…”

“…The antitoxin MqsA is even more unusual because it is the first antitoxin that requires a metal, zinc, for structural stability (19); it is the only E. coli antitoxin that is structured throughout its entire sequence; and it is the first antitoxin demonstrated to bind DNA via its C-terminal and not N-terminal domain (19, 27, 36 -39). Critically, in addition to binding its own promoter (19,34,35,40), MqsA and the MqsR⅐MqsA complex are also the only known antitoxin/TA pair that bind not only their own promoter but also the promoters of other genes that play important roles in E. coli physiology, including mcbR and spy (19) as well as cspD (31). Moreover, although 14 antitoxin structures have been solved to date, their primary sequences are highly divergent when compared with MqsA (supplemental Table S1) because the sequence identity between MqsA and the most closely related antitoxin, the E. coli protein HigA, is only 13%.…”