Crystal structures ofHpSoj–DNA complexes and the nucleoid-adaptor complex formation in chromosome segregation

Abstract

ParABS, an important DNA partitioning process in chromosome segregation, includes ParA (an ATPase), ParB (a par S binding protein) and parS (a centromere-like DNA). The homologous proteins of ParA and ParB in Helicobacter pylori are Hp Soj and Hp Spo0J, respectively. We analyzed the ATPase activity of Hp Soj and found that it is enhanced by both DNA and Hp Spo0J. Crystal structures of Hp Soj and its DNA complexes revealed a typical ATPase fold and that it is dimeric. DNA binding by Hp Soj is promoted by ATP. The Hp Soj–ATP–DNA complex non-specifically binds DNA through a continuous basic binding patch formed by lysine residues, with a single DNA-binding site. This complex exhibits a DNA-binding adept state with an active ATP-bound conformation, whereas the Hp Soj–ADP–DNA complex may represent a transient DNA-bound state. Based on structural comparisons, Hp Soj exhibits a similar DNA binding surface to the bacterial ParA superfamily, but the archaeal ParA superfamily exhibits distinct non-specific DNA-binding via two DNA-binding sites. We detected the Hp Spo0J– Hp Soj–DNA complex by electron microscopy and show that this nucleoid-adaptor complex (NAC) is formed through Hp Soj and Hp Spo0J interaction and parS DNA binding. NAC formation is promoted by Hp Soj participation and specific parS DNA facilitation.

Molecular architecture of the DNA‐binding sites of the P‐loop ATPases MipZ and ParA fromCaulobacter crescentus

“…S9). In line with previous work (35,37), these findings demonstrate that the DNA-binding activity of ParA also relies on non-specific electrostatic interactions with the DNA phosphate backbone. Interestingly, however, only one of the residues identified in ParA (K233) has an equivalent in the DNA-binding pocket of MipZ (R242), because the DNA-interacting region of MipZ is largely part an insertion that is not conserved in other P-loop ATPases (Fig.…”

“…Notably, the crystal structure of the H. pylori ParA-DNA complex revealed a conserved leucine residue that is in direct contact with the DNA molecule. However, this interaction involves the main chain amide group rather than the hydrophobic side chain (37). A similar situation may be observed for MipZ, with mutations in L237 and L248 leading to distortions in the main chain that impede its interaction with the phosphate backbone.…”

“…Our knowledge of the determinants mediating the DNA-binding activity of P-loop ATPases is largely based on studies of canonical ParA homologs. For this group of proteins, two distinct modes of DNA binding, mediated by different regions of the dimer surface, have been reported (37,38). Moreover, the residues identified in previous studies are not universally conserved among DNA-binding members of the ParA-like ATPase family.…”

“…Previously, two arginine residues at the dimer interface of ParA were shown to be critical for DNA binding (28,35). Moreover, a recent crystallographic study has suggested that HpParA interacts with DNA in a region homologous to the DNA-binding interface of MipZ (37). However, a comprehensive analysis of the determinants that mediate the ParA-DNA interaction in solution is still missing.…”

“…Residues homologous to these arginines were later also implicated in the DNA-binding activity of PpfA from Rhodobacter sphaeroides and PomZ from Myxococcus xanthus (17,20). More evidence for the importance of positively charged amino acids has recently come from the crystal structures of ParA-DNA complexes formed by ParA homologs from Helicobacter pylori (HpParA) (37) and Sulfolobus solfataricus pNOB8 (38), which each identified several surface-exposed lysine residues that are in direct contact with the DNA ligand. However, the binding interface pNOB8 ParA was different from that of its H. pylori homolog or R. capsulatus PpfA (38), suggesting that ParA proteins could have evolved different modes of DNA binding.…”