Related Experiment Video

Updated: Jul 11, 2026

FLIM-FRET Measurements of Protein-Protein Interactions in Live Bacteria.
09:26

FLIM-FRET Measurements of Protein-Protein Interactions in Live Bacteria.

Published on: August 25, 2020

Ciprofloxacin interactions with bacterial protein OmpF: modelling of FRET from a multi-tryptophan protein trimer

Fábio Fernandes1, Patrícia Neves, Paula Gameiro

  • 1Centro de Química-Física Molecular, Instituto Superior Técnico, Lisbon, Portugal.

Biochimica Et Biophysica Acta
|September 29, 2007
PubMed

Insights

This study quantifies ciprofloxacin antibiotic binding to bacterial outer membrane protein F (OmpF) using Förster resonance energy transfer (FRET). The binding constant (K(B)) was determined, offering insights into antibiotic uptake mechanisms.

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Microbiology

Background:

  • Outer membrane protein F (OmpF) facilitates fluoroquinolone antibiotic uptake in bacteria.
  • Understanding antibiotic-protein interactions is crucial for combating bacterial resistance.

Purpose of the Study:

  • To quantify the binding affinity of ciprofloxacin to OmpF within a lipid membrane.
  • To develop and apply a Förster resonance energy transfer (FRET) methodology for analyzing complex protein-ligand interactions.

Main Methods:

  • Utilized Förster resonance energy transfer (FRET) to measure ciprofloxacin-OmpF binding.
  • Developed specific FRET formalisms to account for multiple tryptophan residues in OmpF.
  • Analyzed fluorescence quenching to determine binding constants.

Main Results:

  • Determined upper and lower boundaries for the ciprofloxacin-OmpF binding constant (K(B)).
  • Binding constants varied based on the assumed ciprofloxacin binding site location within OmpF.
  • log (K(B)) ranged from 3.15-3.62 or 3.58-4.00.

Conclusions:

  • The developed FRET methodology accurately quantifies antibiotic binding to outer membrane proteins.
  • The findings provide insights into the molecular basis of fluoroquinolone uptake by bacteria.
  • This approach is adaptable for studying similar protein-ligand interactions in various geometries.

More Related Videos

Imaging Protein-protein Interactions in vivo11:15

Imaging Protein-protein Interactions in vivo

Published on: October 10, 2010

Visualization of Bacterial Resistance using Fluorescent Antibiotic Probes08:23

Visualization of Bacterial Resistance using Fluorescent Antibiotic Probes

Published on: March 2, 2020

Related Concept Videos

Inhibitors of Bacterial DNA Synthesis01:28

Inhibitors of Bacterial DNA Synthesis

Bacterial pathogens depend on precise and efficient DNA replication to sustain infection. Two type II topoisomerases—DNA gyrase and topoisomerase IV—are critical to this process, as they resolve DNA supercoiling and unlink chromosomes during replication. Fluoroquinolones, synthetic derivatives of quinolones, exploit this mechanism by stabilizing the transient DNA–enzyme cleavage complex, preventing strand religation, and causing lethal double-strand breaks. These antibiotics are selectively...
Repressible Operon: trp Operon01:21

Repressible Operon: trp Operon

The trp operon in Escherichia coli exemplifies a repressible operon. It regulates the synthesis of tryptophan through repressor-mediated transcriptional control and attenuation. This dual regulatory mechanism ensures tryptophan biosynthesis occurs only when needed, conserving cellular resources.Structure of the trp OperonThe trp operon consists of five structural genes (trpE, trpD, trpC, trpB, and trpA) that encode enzymes for tryptophan biosynthesis. These genes are transcribed as a single...
Protein-protein Interfaces02:04

Protein-protein Interfaces

Many proteins form complexes to carry out their functions, making protein-protein interactions (PPIs) essential for an organism's survival. Most PPIs are stabilized by numerous weak noncovalent chemical forces. The physical shape of the interfaces determines the way two proteins interact. Many globular proteins have closely-matching shapes on their surfaces, which form a large number of weak bonds. Additionally, many PPIs occur between two helices or between a surface cleft and a polypeptide...
Protein Complexes with Interchangeable Parts01:57

Protein Complexes with Interchangeable Parts

Groups of proteins may form a complex where each protein in this complex has a different role in the overall execution of the complex’s function. Often some of the proteins in the complex can be replaced by a closely related variant to give a complex that contains many of the same components yet is functionally distinct.
The SCF ubiquitin ligase is a protein complex of five individual proteins. This complex attaches ubiquitin to other target proteins to mark them for degradation. In order to...
Inhibitors of Bacterial Protein Synthesis01:25

Inhibitors of Bacterial Protein Synthesis

Aminoglycosides constitute a highly potent class of bactericidal antibiotics that exert their antimicrobial effects by targeting the bacterial ribosome, specifically disrupting protein synthesis. These polycationic molecules consist of amino-modified sugars linked via glycosidic bonds to an aminocyclitol core such as 2-deoxystreptamine or streptamine. Their strong positive charges facilitate tight binding to the negatively charged phosphate backbone of ribosomal RNA (rRNA), primarily at the 16S...
Cooperative Allosteric Transitions01:58

Cooperative Allosteric Transitions

Cooperative allosteric transitions can occur in multimeric proteins, where each subunit of the protein has its own ligand-binding site. When a ligand binds to any of these subunits, it triggers a conformational change that affects the binding sites in the other subunits; this can change the affinity of the other sites for their respective ligands. The ability of the protein to change the shape of its binding site is attributed to the presence of a mix of flexible and stable segments in the...