3 research outputs found
Bicarbonate Alters Bacterial Susceptibility to Antibiotics by Targeting the Proton Motive Force
The
antibacterial properties of sodium bicarbonate have been known for
years, yet the molecular understanding of its mechanism of action
is still lacking. Utilizing chemical–chemical combinations,
we first explored the effect of bicarbonate on the activity of conventional
antibiotics to infer on the mechanism. Remarkably, the activity of
8 classes of antibiotics differed in the presence of this ubiquitous
buffer. These interactions and a study of mechanism of action revealed
that, at physiological concentrations, bicarbonate is a selective
dissipater of the pH gradient of the proton motive force across the
cytoplasmic membrane of both Gram-negative and Gram-positive bacteria.
Further, while components that make up innate immunity have been extensively
studied, a link to bicarbonate, the dominant buffer in the extracellular
fluid, has never been made. Here, we also explored the effects of
bicarbonate on components of innate immunity. Although the immune
response and the buffering system have distinct functions in the body,
we posit there is interplay between these, as the antimicrobial properties
of several components of innate immunity were enhanced by a physiological
concentration of bicarbonate. Our findings implicate bicarbonate as
an overlooked potentiator of host immunity in the defense against
pathogens. Overall, the unique mechanism of action of bicarbonate
has far-reaching and predictable effects on the activity of innate
immune components and antibiotics. We conclude that bicarbonate has
remarkable power as an antibiotic adjuvant and suggest that there
is great potential to exploit this activity in the discovery and development
of new antibacterial drugs by leveraging testing paradigms that better
reflect the physiological concentration of bicarbonate
Exploiting the Sensitivity of Nutrient Transporter Deletion Strains in Discovery of Natural Product Antimetabolites
Actinomycete
secondary metabolites are a renowned source of antibacterial chemical
scaffolds. Herein, we present a target-specific approach that increases
the detection of antimetabolites from natural sources by screening
actinomycete-derived extracts against nutrient transporter deletion
strains. On the basis of the growth rescue patterns of a collection
of 22 <i>Escherichia coli</i> (<i>E. coli</i>)
auxotrophic deletion strains representative of the major nutrient
biosynthetic pathways, we demonstrate that antimetabolite detection
from actinomycete-derived extracts prepared using traditional extraction
platforms is masked by nutrient supplementation. In particular, we
find poor sensitivity for the detection of antimetabolites targeting
vitamin biosynthesis. To circumvent this and as a proof of principle,
we exploit the differential activity of actinomycete extracts against <i>E. coli ΔyigM</i>, a biotin transporter deletion strain
versus wildtype <i>E. coli</i>. We achieve more than a 100-fold
increase in antimetabolite sensitivity using this method and demonstrate
a successful bioassay-guided purification of the known biotin antimetabolite,
amiclenomycin. Our findings provide a unique solution to uncover the
full potential of naturally derived antibiotics
Structural and Kinetic Characterization of Diazabicyclooctanes as Dual Inhibitors of Both Serine-β-Lactamases and Penicillin-Binding Proteins
Avibactam
is a diazabicyclooctane β-lactamase inhibitor possessing
outstanding but incomplete efficacy against multidrug-resistant Gram-negative
pathogens in combination with β-lactam antibiotics. Significant
pharmaceutical investment in generating derivatives of avibactam warrants
a thorough characterization of their activity. We show here through
structural and kinetic analysis that select diazabicyclooctane derivatives
display effective but varied inhibition of two clinically important
β-lactamases (CTX-M-15 and OXA-48). Furthermore, these derivatives
exhibit considerable antimicrobial activity (MIC ≤ 2 μg/mL)
against clinical isolates of <i>Pseudomonas aeruginosa</i>, <i>Escherichia coli</i>, and <i>Enterobacter spp</i>. Imaging of cell phenotype along with structural and biochemical
experiments unambiguously demonstrate that this activity, in <i>E. coli</i>, is a result of targeting penicillin-binding protein
2. Our results suggest that structure–activity relationship
studies for the purpose of drug discovery must consider both β-lactamases
and penicillin-binding proteins as targets. We believe that this approach
will yield next-generation combination or monotherapies with an expanded
spectrum of activity against currently untreatable Gram-negative pathogens