Biomedical subjects

Daniela Barillà

Publications and source records attributed to Daniela Barillà.

7 recordsLinked to original sources

PubMed2006

Assembling the bacterial segrosome.

Genome segregation in prokaryotes is a highly ordered process that integrates with DNA replication, cytokinesis and other fundamental facets of the bacterial cell cycle. The segrosome is the nucleoprotein complex that mediates DNA segregation in bacteria, its assembly and organization is best understood for plasmid partition. The recent elucidation of structures of the ParB plasmid segregation protein bound to centromeric DNA, and of the tertiary structures of other segregation proteins, are key milestones in the path to deciphering the molecular basis of bacterial DNA segregation.

Centromere↗

PubMed2006

The bacterial segrosome: a dynamic nucleoprotein machine for DNA trafficking and segregation.

The genomes of unicellular and multicellular organisms must be partitioned equitably in coordination with cytokinesis to ensure faithful transmission of duplicated genetic material to daughter cells. Bacteria use sophisticated molecular mechanisms to guarantee accurate segregation of both plasmids and chromosomes at cell division. Plasmid segregation is most commonly mediated by a Walker-type ATPase and one of many DNA-binding proteins that assemble on a cis-acting centromere to form a nucleoprotein complex (the segrosome) that mediates intracellular plasmid transport. Bacterial chromosome segregation involves a multipartite strategy in which several discrete protein complexes potentially participate. Shedding light on the basis of genome segregation in bacteria could indicate new strategies aimed at combating pathogenic and antibiotic-resistant bacteria.

Amino Acid Sequence↗

PubMed2005

The unstructured N-terminal tail of ParG modulates assembly of a quaternary nucleoprotein complex in transcription repression.

ParG is the prototype of a group of small (<10 kDa) proteins involved in accurate plasmid segregation. The protein is a dimeric DNA binding factor, which consists of symmetric paired C-terminal domains that interleave into a ribbon-helix-helix fold that is crucial for the interaction with DNA, and unstructured N-terminal domains of previously unknown function. Here the ParG protein is shown to be a transcriptional repressor of the parFG genes. The protein assembles on its operator site initially as a tetramer (dimer of dimers) and, at elevated protein concentrations, as a pair of tetramers. Progressive deletion of the mobile N-terminal tails concomitantly decreased transcriptional repression by ParG and perturbed the DNA binding kinetics of the protein. The flexible tails are not necessary for ParG dimerization but instead modulate the organization of a higher order nucleoprotein complex that is crucial for proper transcriptional repression. This is achieved by transient associations between the flexible and folded domains in complex with the target DNA. Numerous ParG homologs encoded by plasmids of Gram-negative bacteria similarly are predicted to possess N-terminal disordered tails, suggesting that this is a common feature of partition operon autoregulation. The results provide new insights into the role of natively unfolded domains in protein function, the molecular mechanisms of transcription regulation, and the control of plasmid segregation.

Base Sequence↗

PubMed2005

Bacterial DNA segregation dynamics mediated by the polymerizing protein ParF.

Prokaryotic DNA segregation most commonly involves members of the Walker-type ParA superfamily. Here we show that the ParF partition protein specified by the TP228 plasmid is a ParA ATPase that assembles into extensive filaments in vitro. Polymerization is potentiated by ATP binding and does not require nucleotide hydrolysis. Analysis of mutations in conserved residues of the Walker A motif established a functional coupling between filament dynamics and DNA partitioning. The partner partition protein ParG plays two separable roles in the ParF polymerization process. ParF is unrelated to prokaryotic polymerizing proteins of the actin or tubulin families, but is a homologue of the MinD cell division protein, which also assembles into filaments. The ultrastructures of the ParF and MinD polymers are remarkably similar. This points to an evolutionary parallel between DNA segregation and cytokinesis in prokaryotic cells, and reveals a potential molecular mechanism for plasmid and chromosome segregation mediated by the ubiquitous ParA-type proteins.

1-Acylglycerol-3-Phosphate O-Acyltransferase↗

PubMed2005

Protein diversity confers specificity in plasmid segregation.

The ParG segregation protein (8.6 kDa) of multidrug resistance plasmid TP228 is a homodimeric DNA-binding factor. The ParG dimer consists of intertwined C-terminal domains that adopt a ribbon-helix-helix architecture and a pair of flexible, unstructured N-terminal tails. A variety of plasmids possess partition loci with similar organizations to that of TP228, but instead of ParG homologs, these plasmids specify a diversity of unrelated, but similarly sized, partition proteins. These include the proteobacterial pTAR, pVT745, and pB171 plasmids. The ParG analogs of these plasmids were characterized in parallel with the ParG homolog encoded by the pseudomonal plasmid pVS1. Like ParG, the four proteins are dimeric. No heterodimerization was detectable in vivo among the proteins nor with the prototypical ParG protein, suggesting that monomer-monomer interactions are specific among the five proteins. Nevertheless, as with ParG, the ParG analogs all possess significant amounts of unordered amino acid residues, potentially highlighting a common structural link among the proteins. Furthermore, the ParG analogs bind specifically to the DNA regions located upstream of their homologous parF-like genes. These nucleoprotein interactions are largely restricted to cognate protein-DNA pairs. The results reveal that the partition complexes of these and related plasmids have recruited disparate DNA-binding factors that provide a layer of specificity to the macromolecular interactions that mediate plasmid segregation.

Bacterial Proteins↗

PubMed2003

Architecture of the ParF*ParG protein complex involved in prokaryotic DNA segregation.

The mechanism by which low copy number plasmids are segregated at cell division involves the concerted action of two plasmid-encoded proteins that assemble on a centromere-like site. This study explores the topology of the DNA segregation machinery specified by the parFG locus of TP228, a partition system which is phylogenetically distinct from more well-characterized archetypes. A variety of genetic, biochemical and biophysical strategies revealed that the ParG protein is dimeric. ParF, which is more closely related to the cell division regulator MinD than to the prototypical ParA partition protein of plasmid P1, is instead multimeric and its polymeric state appears to be modulated by ATP which correlates with the proposed ATP-binding activity of ParF. ParG interacts in a sequence-specific manner with the DNA region upstream of the parFG locus and this binding is modulated by ParF. Intriguingly, the ParF and ParG proteins form at least two types of discrete complex in the absence of this region suggesting that the assembly dynamics of these proteins onto DNA is intricate.

Adenosine Triphosphate↗

PubMed2003

ParG, a protein required for active partition of bacterial plasmids, has a dimeric ribbon-helix-helix structure.

The ParG protein (8.6 kDa) is an essential component of the DNA partition complex of multidrug resistance plasmid TP228. ParG is a dimer in solution, interacts with DNA sequences upstream of the parFG genes and also with the ParF partition protein both in the absence and presence of target DNA. Here, the solution nuclear magnetic resonance structure of ParG is reported. The ParG dimer is composed of a folded domain formed by two closely intertwined C-terminal parts (residues 33-76), and two highly mobile tails consisting of N-terminal regions (residues 1-32). The folded part of ParG has the ribbon-helix-helix (RHH) architecture similar to that of the Arc/MetJ superfamily of DNA-binding transcriptional repressors, although the primary sequence similarity is very low. ParG interacts with DNA predominantly via its folded domain; this interaction is coupled with ParG oligomerization. The dimeric RHH structure of ParG suggests that it binds to DNA by inserting the double-stranded beta-sheet into the major groove of DNA, in a manner similar to transcriptional repressors from the Arc/MetJ superfamily, and that ParG can function as a transcriptional repressor itself. A new classification of proteins belonging to the Arc/MetJ superfamily and ParG homologues is proposed, based on the location of a conserved positively charged residue at either the beginning or at the end of the beta-strand which forms part of the DNA recognition motif.

Amino Acid Sequence↗