The five-to-six-coordination transition of ferric human serum heme-albumin is allosterically-modulated by ibuprofen and warfarin: a combined XAS and MD study.

Carlo Meneghini,Paolo Ascenzi,Gabriella Fanali,Loris Leboffe,Settimio Mobilio,Mauro Fasano,Giorgio Colombo,Monica Bionducci,Massimiliano Meli,Rizwan H Khan

doi:10.1371/journal.pone.0104231

Abstract

Human serum albumin (HSA) is involved physiologically in heme scavenging; in turn, heme-albumin (HSA-heme-Fe) displays globin-like properties. Here, the allosteric effect of ibuprofen and warfarin on the local atomic structure around the ferric heme-Fe (heme-Fe(III)) atom of HSA-heme-Fe (HSA-heme-Fe(III)) has been probed by Fe-K edge X-ray absorption spectroscopy (XAS). The quantitative analysis of the Fe-K edge extended X-ray absorption fine structure (EXAFS) signals and modeling of the near edge (XANES) spectral features demonstrated that warfarin and ibuprofen binding modify the local structure of the heme-Fe(III). Combined XAS data analysis and targeted molecular dynamics (MD) simulations provided atomic resolution insights of protein structural rearrangements required to accommodate the heme-Fe(III) upon ibuprofen and warfarin binding. In the absence of drugs, the heme-Fe(III) atom is penta-coordinated having distorted 4+1 configuration made by the nitrogen atoms of the porphyrin ring and the oxygen phenoxy atom of the Tyr161 residue. MD simulations show that ibuprofen and warfarin association to the secondary fatty acid (FA) binding site 2 (FA2) induces a reorientation of domain I of HSA-heme-Fe(III), this leads to the redirection of the His146 residue providing an additional bond to the heme-Fe(III) atom, providing the 5+1 configuration. The comparison of Fe-K edge XANES spectra calculated using MD structures with those obtained experimentally confirms the reliability of the proposed structural model. As a whole, combining XAS and MD simulations it has been possible to provide a reliable model of the heme-Fe(III) atom coordination state and to understand the complex allosteric transition occurring in HSA-heme-Fe(III) upon ibuprofen and warfarin binding.

Highlights

Human serum albumin (HSA), the most abundant protein in plasma, represents the main determinant of plasma oncotic pressure, is the major modulator of fluid distribution within the body compartments, and displays an extraordinary ligand-binding capacity [1,2]
Fe-K edge (EFe = 7.112 keV) X-ray absorption spectra of HSAheme-Fe(III) in the absence and presence of ibuprofen or warfarin were collected at the BM23 XAFS beamline
The edge position is weakly affected by addition of ibuprofen and warfarin binding to HSA-heme-Fe(III), demonstrating that drugs do not affect the heme-Fe(III) atom electronic state, but mainly modify the atomic structure around it

Summary

Introduction

Human serum albumin (HSA), the most abundant protein in plasma (ca. 761024 M), represents the main determinant of plasma oncotic pressure, is the major modulator of fluid distribution within the body compartments, and displays an extraordinary ligand-binding capacity [1,2]. Human serum albumin (HSA), the most abundant protein in plasma 761024 M), represents the main determinant of plasma oncotic pressure, is the major modulator of fluid distribution within the body compartments, and displays an extraordinary ligand-binding capacity [1,2]. HSA provides a depot and carrier for many endogenous and exogenous compounds, affects pharmacokinetics of many drugs, induces the metabolic modification(s) of some ligands, renders potential toxins harmless, accounts for most of the anti-oxidant capacity of human plasma, and displays (pseudo-)enzymatic properties [1,2]. HSA is a single non-glycosylated all-a chain protein, constituted by 585 amino acids, containing three homologous domains (labeled I, II, and III). HSA displays at least nine fatty acid (FA) binding clefts (FA1–FA9) [6]. Ibuprofen, a non-steroidal anti-inflammatory drug, and warfarin,

Methods

Results

Conclusion