Bismuth-based drugs inhibit Porphyromonas gingivalis: proteomic/transcriptomic profiling & molecular mechanisms

Abstract

Backgrounds: Porphyromonas gingivalis is a ‘keystone’ pathogen for the highly common periodontal disease worldwide. Objectives: This study aimed to investigate the protein targets and effects of bismuth-based drugs on the gene expression of P. gingivalis, through metal-based proteomics and RNA seq & Real-time PCR. Methods: The minimal inhibition concentration of ranitidine bismuth citrate (RBC) for P. gingivalis was determined after 48-hour incubation with series-diluted RBC and other antimicrobials. Fluorescence-labeling-based 2D-PAGE was performed on P. gingivalis cell lysates with a fluorescence probe of Bi3+-TRACER. The identified proteins were selected and cloned, and subsequently recombinant proteins were overexpressed and purified for activity determination. The effects of bismuth on mRNA expression profile of P. gingivalis were analyzed by RNA seq, and selected genes were further evaluated by Real-time PCR. Conclusions: It was shown that bismuth could inhibit P. gingivalis growth, and enhance the bacterial sensitivity to antimicrobials (e.g. metronidazole and chlorhexidine). Multiple protein spots on the 2D-PAGE of P. gingivalis lysates were lit up by Bi3+-TRACER after UV-radiation, and dozens of potential bismuth-associating proteins were identified by peptide mass fingerprinting on these lit-up spots. Notably, bismuth could partially or even completely inactivate these selected proteins. Whole-genome RNA seq revealed that the expression of substantial amounts of genes with various functions was markedly affected during the first hour of bismuth treatment which was selectively verified by Real-time PCR. This pioneering study indicates that P. gingivalis undergoes extensive influences by bismuth which leads to bacterial inhibition.