Three ParA Dimers Cooperatively Assemble on Type Ia Partition Promoters.

François Boudsocq,Jean-Yves Bouet,Sophie Barbe,Maya Salhi

doi:10.3390/genes12091345

Abstract

Accurate DNA segregation is essential for faithful inheritance of genetic material. In bacteria, this process is mainly ensured by partition systems composed of two proteins, ParA and ParB, and a centromere site. Auto-regulation of Par operon expression is important for efficient partitioning and is primarily mediated by ParA for type Ia plasmid partition systems. For the F-plasmid, four ParAF monomers were proposed to bind to four repeated sequences in the promoter region. By contrast, using quantitative surface-plasmon-resonance, we showed that three ParAF dimers bind to this region. We uncovered that one perfect inverted repeat (IR) motif, consisting of two hexamer sequences spaced by 28-bp, constitutes the primary ParAF DNA binding site. A similar but degenerated motif overlaps the former. ParAF binding to these motifs is well supported by biochemical and modeling analyses. Molecular dynamics simulations predict that the winged-HTH domain displays high flexibility, which may favor the cooperative ParA binding to the promoter. We propose that three ParAF dimers bind cooperatively to overlapping motifs, thus covering the promoter region. A similar organization is found on closely related and distant plasmid partition systems, suggesting that such promoter organization for auto-regulated Par operons is widespread and may have evolved from a common ancestor.

Highlights

Fdimer on the promoter region, we proposed that two hexamer motifs form a pera ParAF dimer on the promoter region, we proposed that two hexamer motifs form a fect inverted repeat (IR)
To investigate how ParAF interacts with its promoter, we performed a combination of three techniques measuring protein-DNA interactions
No Surface Plasmon Resonance (SPR) signal was detected if PparABF DNA was replaced by non-specific DNA probes of the same length, indicating that ParAF binding to the promoter region is highly specific in the absence (Figure 2B) or presence of ADP (Figure S1C)

Summary

Introduction

Stable inheritance of low-copy-number bacterial replicons (chromosomes and plasmids) requires an active partition mechanism. Each of such replicons typically carries a selfspecific partition module comprising a two-gene operon and an adjacent set of short sequence repeats that acts as a centromere. The first gene encodes an ATPase and the second a centromere-binding protein. Partition systems have been classified into three main classes depending on the type of ATPase encoded with Type I, II and III, which are characterized by deviant Walker A, actin-like and tubulin-like NTPases, respectively (for review [1,2]). Type I systems, generically termed ParABS, are the most common type in the bacterial world, being widespread on low-copy-number plasmids and the only type present on chromosomes.

Methods

Results

Conclusion