Introduction to the 6th Biometals Webinars, Metals, metabolites, and the variable host environment, Mechanistic studies of the bacterial nitrosative stress sensor NsrR

Mechanistic studies of the bacterial nitrosative stress sensor NsrR Nitric oxide (NO) is a ubiquitous signalling molecule and cytotoxic agent. In order to survive and thrive, organisms must be able to respond to NO, and the regulatory protein NsrR, a member of the Rrf2 protein superfamily, plays a major role in this as it is found in a wide range of benign and pathogenic bacteria. NsrR binds a [4Fe-4S] cluster as a cofactor and, in the absence of NO, represses transcription of genes primarily involved NO detoxification. Previously, we have shown that [4Fe-4S] NsrR reacts rapidly with NO, in a complex reaction involving multiple NO molecules per cluster, resulting in formation of NsrR-bound iron-nitrosyl species [3]. Here, we sought to investigate the reaction of [4Fe-4S] NsrR bound to DNA, in order to determine when high affinity DNA-binding is lost. Using surface plasmon resonance (SPR) and native mass spectrometry (MS) NsrR binding to the promoter regions of two NsrR-regulated genes, hmpA1 and hmpA2 was probed, revealing the nature of binding and the reaction of DNA-bound NsrR with NO in remarkable detail.