Defined DNA sequences promote the assembly of a bacterial protein into distinct amyloid nanostructures.
RepA, the replication initiator protein of Pseudomonas pPS10 plasmid, is made of two winged-helix (WH) domains. RepA dimers undergo a structural transformation upon binding to origin DNA sequences (iterons), resulting in monomerization and alpha-helix into beta-strand conversion. This affects the N-terminal domain (WH1) and generates a metastable intermediate. Here it is shown that the interaction of short dsDNA oligonucleotides, including iteron or operator RepA targets, with the isolated WH1 domain promotes the assembly of different nanostructures. These range from irregular aggregates to amyloid spheroids and fibers. Their intrinsic order inversely correlates with the extent of the transformation induced by each DNA sequence on RepA. However, DNA is not a constituent of the assembled fibers, in agreement with the protein-only principle for amyloid structure. Thus, the RepA-WH1 domain on DNA binding mimics the behavior of the mammalian prion protein. The stretch of amino acids responsible for WH1 aggregation has been identified, leading to the design of mutants with enhanced or reduced amyloidogenicity and the synthesis of a peptide that assembles into a cross-beta structure. RepA amyloid assemblies could have a role in the negative regulation of plasmid replication. This article underlines the potential of specific nucleic acid sequences in promoting protein amyloidogenesis at nearly physiological conditions.
Giraldo
Rafael
R
Department of Molecular Microbiology, Centro de Investigaciones Biológicas, Consejo Superior de Investigaciones Científicas, C/Ramiro de Maeztu 9, 28040 Madrid, Spain. rgiraldo@cib.csic.es
D000595:Amino Acid Sequence; D000682:Amyloid / Q000737:chemistry / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure*; D001483:Base Sequence; D002942:Circular Dichroism; D004247:DNA / Q000235:genetics*; D046529:Microscopy, Electron, Transmission; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D049329:Nanostructures / Q000737:chemistry / Q000648:ultrastructure*; D017434:Protein Structure, Tertiary; D011549:Pseudomonas / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure*
D000682:Amyloid; D014157:Transcription Factors; D004247:DNA