Characterization of Two Distinct Modes of Drug Binding to Human Intestinal Fatty Acid Binding Protein

The aqueous cytoplasm of cells poses a potentially significant barrier for many lipophilic drugs to reach their sites of action. Fatty acid binding proteins (FABPs) bind to poorly water-soluble fatty acids (FAs) and lipophilic compounds and facilitate their intracellular transport. Several structures of FA in complex with FABPs have been described, but data describing the binding sites of other lipophilic ligands including drugs are limited. Here the environmentally sensitive fluorophores, 1-anilinonapthalene 8-sulfonic acid (ANS), and 11-dansylamino undecanoic acid (DAUDA) were used to investigate drug binding to human intestinal FABP (hIFABP). Most drugs that bound hIFABP were able to displace both ANS and DAUDA. A notable exception was ketorolac, a non-steroidal anti-inflammatory drug that bound to hIFABP and displaced DAUDA but failed to displace ANS. Isothermal titration calorimetry revealed that for the majority of ligands including FA, ANS, and DAUDA, binding to hIFABP was exothermic. In contrast, ketorolac binding to hIFABP was endothermic and entropy-driven. The X-ray crystal structure of DAUDA–hIFABP revealed a FA-like binding mode where the carboxylate of DAUDA formed a network of hydrogen bonds with residues at the bottom of the binding cavity and the dansyl group interacted with residues in the portal region. In contrast, NMR chemical shift perturbation (CSP) data suggested that ANS bound only toward the bottom of the hIFABP cavity, whereas ketorolac occupied only the portal region. The CSP data further suggested that ANS and ketorolac were able to bind simultaneously to hIFABP, consistent with the lack of displacement of ANS observed by fluorescence and supported by a model of the ternary complex. The NMR solution structure of the ketorolac–hIFABP complex therefore describes a newly characterized, hydrophobic ligand binding site in the portal region of hIFABP

Meteorological conditions.

Features describing dispersive and non-dispersive conditions.</p

ERA5-Land reanalysis.

Meteo variables used in this study and link to the ERA5-Land Reanalysis for the period January 1—June 30 (2016–2020). (DOCX)</p

Percentage concentration differences of pollutants under different conditions.

Percentage changes of PM10 (blue), PM2.5 (red), NO2 (green), and O3 (yellow) of daily mean concentrations. Darker to lighter: dispersive pre-lockdown, dispersive lockdown, dispersive post-lockdown, non-dispersive pre-lockdown, non-dispersive lockdown, non-dispersive post-lockdown. (PDF)</p

Concentration data availability.

Green cells represent concentration data of PM10, PM 2.5, NO2 and O3 available for the individual cities; red cells represent a lack of data. (DOCX)</p

Oxford Stringency Index.

Evolution of the Oxford Stringency Index in selected European countries.</p

Concentration data.

Links to PM10, PM2.5, NO2 and O3 concentration data repository for the period January 1—June 30 (2016–2020). (DOCX)</p

Study domain.

Map showing the cities examined in the study.</p

Meteorological classes.

Frequencies of occurrence (%) of meteorological classes in 2016–2019 and 2020, during the lockdown and no-lockdown periods. (PDF)</p

Oxford Stringency Index.

OxSI (January 1—June 30, 2020) link to data source. (DOCX)</p