The CTPase activity of ParB determines the size and dynamics of prokaryotic DNA partition complexes

Manuel Osorio-Valeriano,Florian Altegoer,Chandan K Das,Wieland Steinchen,Gaël Panis,Lara Connolley,Giacomo Giacomelli,Helge Feddersen,Laura Corrales-Guerrero,Pietro I Giammarinaro,Juri Hanßmann,Marc Bramkamp,Patrick H Viollier,Seán Murray,Lars V Schäfer,Gert Bange,Martin Thanbichler

doi:10.1016/j.molcel.2021.09.004

Abstract

ParB-like CTPases mediate the segregation of bacterial chromosomes and low-copy number plasmids. They act as DNA-sliding clamps that are loaded at parS motifs in the centromere of target DNA molecules and spread laterally to form large nucleoprotein complexes serving as docking points for the DNA segregation machinery. Here, we solve crystal structures of ParB in the pre- and post-hydrolysis state and illuminate the catalytic mechanism of nucleotide hydrolysis. Moreover, we identify conformational changes that underlie the CTP- and parS-dependent closure of ParB clamps. The study of CTPase-deficient ParB variants reveals that CTP hydrolysis serves to limit the sliding time of ParB clamps and thus drives the establishment of a well-defined ParB diffusion gradient across the centromere whose dynamics are critical for DNA segregation. These findings clarify the role of the ParB CTPase cycle in partition complex assembly and function and thus advance our understanding of this prototypic CTP-dependent molecular switch.

Highlights

The ParB family of proteins has emerged as a class of molecular switches that require CTP for proper function (Osorio-Valeriano et al, 2019; Soh et al, 2019)
A second study clarified the structure of a homologous CTP-bound nucleotide-binding domain (NBD) from PadC (Osorio-Valeriano et al, 2019), an accessory protein of the ParABS system in M. xanthus (Lin et al, 2017)
Consistent with the crystallographic data (Figure S2C), the simulations show that the loss of E93 increases the flexibility of Q52 and R97 and destabilizes loop a1/a2 in the region surrounding F57 (Figure S2E). These results show that the catalytic residues Q52 and E93 have a critical role in the nucleotidedependent conformational dynamics of ParB proteins

Summary

Introduction

The ParB family of proteins has emerged as a class of molecular switches that require CTP for proper function (Osorio-Valeriano et al, 2019; Soh et al, 2019). After initial specific interaction with individual parS sites, it spreads into adjacent DNA segments, giving rise to a large nucleoprotein assembly (partition complex) that typically includes 10–20 kb of the origin region (Breier and Grossman, 2007; Lynch and Wang, 1995; Murray et al, 2006; Rodionov et al, 1999). In addition to its role in origin segregation, the partition complex can have several additional functions (Kawalek et al, 2020), including the polar attachment of the origin regions (Bowman et al, 2008; Donovan et al, 2012; Ebersbach et al, 2008; Yamaichi et al, 2012), the loading of bacterial condensin complexes (Bohm et al, 2020; Gruber and Errington, 2009; Sullivan et al, 2009; Tran et al, 2017), and the regulation of division site placement (Thanbichler and Shapiro, 2006; Toro-Nahuelpan et al, 2019)

Results

Discussion

Conclusion