Abstract
Binding of hippuric acid (HA), a uremic toxin, with human serum albumin (HSA) has been examined by isothermal titration calorimetry (ITC), differential scanning calorimetry (DSC), molecular docking, circular dichroism (CD) and fluorescence spectroscopy to understand the reason that govern its impaired elimination through hemodialysis. ITC results shows that the HA binds with HSA at high (K b ∼104) and low affinity (K b ∼103) sites whereas spectroscopic results predict binding at a single site (K b∼103). The HA form complex with HSA that involves electrostatic, hydrogen and hydrophobic binding forces as illustrated by calculated thermodynamic parameters. Molecular docking and displacement studies collectively revealed that HA bound to both site I and site II; however, relatively strongly to the later. Esterase-like activity of HSA confirms the involvement of Arg410 and Tyr411 of Sudlow site II in binding of HA. CD results show slight conformational changes occurs in the protein upon ligation that may be responsible for the discrepancy in van’t Hoff and calorimetric enthalpy change. Furthermore, an increase in and is observed from DSC results that indicate increase in stability of HSA upon binding to HA. The combined results provide that HA binds to HSA and thus its elimination is hindered.
Highlights
Uremic toxins are the compounds which retained in the blood during kidney failure and interact negatively with the normal biological functions of the body [1]
Steady State Fluorescence Quenching Measurements The aromatic fluorophores, tryptophan, tyrosine, and phenylalanine are very sensitive to their microenvironment and used for studying conformational changes associated with drug protein binding
The binding affinity observed by fluorescence spectroscopy took in consideration the location of quencher, fluorophore and so measures local changes around the fluorophores associated with the optical transition [29]
Summary
Uremic toxins are the compounds which retained in the blood during kidney failure and interact negatively with the normal biological functions of the body [1]. Hippuric acid (HA) is one of these compounds that accumulates in the blood, and cause stimulation of ammoniagenesis. It is involved in development of muscular weakness in uremia as it inhibits glucose utilization in muscles [2,3,4]. HA is a compound of pharmacological interest. It is a glycine conjugate of benzoate, which is formed primarily from aromatic amino acids by gastrointestinal flora or may be directly taken as preservatives from food and beverages [7]. Concentration of HA is less than 5 mg/L but increases to values higher than 2476112 mg/L in patients with end-stage renal disease [8]
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