Buforin II-Escherichia coli's DNA interactome: Detailed biophysical characterization revealed nanoscale complexes likely formed by DNA supercoiling
Antimicrobial peptides (AMPs) have emerged as exciting alternatives to the alarming increase of multiresistant bacteria due to their high activity against them through mechanisms that are thought to largely avoid resistance in the long term. Buforin II (BUFII) is an antibacterial peptide hypothesized to kill bacteria by crossing their membranes to interact with intracellular molecules and interrupt key processes for survival. In particular, interactions with DNA have been considered crucial for triggering cell death mechanisms. However, such interactions are still unknown, and thus far, no reports are available describing BUFII-DNA complexes. Here, we describe a complete biophysical study of the interaction between BUFII and Escherichia coli gDNA via spectrofluorimetric, spectroscopic, and microscopic techniques, complemented with whole-genome sequencing. The E. colis DNA-BUFII interactome was isolated by an in vitro pull-down method aided by BUFII-magnetite nanobioconjugates. Our results demonstrated that DNA-BUFII formed round-shape nanoscale complexes by strong electrostatic interactions, likely occurring nonspecifically throughout the entire bacterial genome. Further sequencing of the isolated DNA fragments corroborated this notion and led to hypothesize that BUFII is possibly responsible for inducing DNAs supercoiling. Other evidence for this idea was provided by the significant DNA conformational changes observed upon interaction with BUFII. Even though the evidence found fails to describe the complete action mechanism of BUFII in vivo, our findings pave the way to engineer DNA-peptide supramolecular complexes very precisely, which might find application in the field of gene therapy delivery.