The protein Human Serum Albumin (HSA) is known to undergo conformational transitions towards partially unfolded forms triggered by acidification below pH 4.5. The extent of Fatty Acids (FA) binding has been thought to have an impact on the conformational equilibrium between the native and acid forms and to be a possible explanation for the observation of more than one band in early electrophoretic migration experiments at pH 4. We compared the acid-induced unfolding processes of commercial FA-free HSA, commercial “fatted” HSA and FA-HSA complexes, prepared at FA:HSA molar ratios between 1 and 6 by simple mixing and equilibration. We used a method for continuous acidification based on the hydrolysis of glucono-δ-lactone from pH 7 to pH 2.5, and followed the average protein changes by the blue shift of the intrinsic fluorescence emission and by performing a small angle X-ray scattering analysis on selected samples. The method also allowed for continuous monitoring of the increase of turbidity and laser light scattering of the protein samples related to the release of the insoluble ligands with acidification. Our results showed that the presence of FA interacting with albumin, an aspect often neglected in biophysical studies, affects the conformational response of the protein to acidification, and slightly shifts the loss of the native shape from pH 4.2 to pH 3.6. This effect increased with the FA:HSA molar ratio so that with three molar equivalents a saturation was reached, in agreement with the number of high-affinity binding sites reported for the FA. These findings confirm that a non-uniform level of ligand binding in an albumin sample can be an explanation for the early-observed conformational heterogeneity at pH 4.

The effect of fatty acid binding in the acid isomerizations of albumin investigated with a continuous acidification method / Del Giudice, Alessandra; Galantini, Luciano; Dicko, Cedric; Pavel, Nicolae V.. - In: COLLOIDS AND SURFACES. B, BIOINTERFACES. - ISSN 0927-7765. - STAMPA. - 168:(2018), pp. 109-116. [10.1016/j.colsurfb.2018.03.038]

The effect of fatty acid binding in the acid isomerizations of albumin investigated with a continuous acidification method

Del Giudice, Alessandra
;
Galantini, Luciano;Pavel, Nicolae V.
2018

Abstract

The protein Human Serum Albumin (HSA) is known to undergo conformational transitions towards partially unfolded forms triggered by acidification below pH 4.5. The extent of Fatty Acids (FA) binding has been thought to have an impact on the conformational equilibrium between the native and acid forms and to be a possible explanation for the observation of more than one band in early electrophoretic migration experiments at pH 4. We compared the acid-induced unfolding processes of commercial FA-free HSA, commercial “fatted” HSA and FA-HSA complexes, prepared at FA:HSA molar ratios between 1 and 6 by simple mixing and equilibration. We used a method for continuous acidification based on the hydrolysis of glucono-δ-lactone from pH 7 to pH 2.5, and followed the average protein changes by the blue shift of the intrinsic fluorescence emission and by performing a small angle X-ray scattering analysis on selected samples. The method also allowed for continuous monitoring of the increase of turbidity and laser light scattering of the protein samples related to the release of the insoluble ligands with acidification. Our results showed that the presence of FA interacting with albumin, an aspect often neglected in biophysical studies, affects the conformational response of the protein to acidification, and slightly shifts the loss of the native shape from pH 4.2 to pH 3.6. This effect increased with the FA:HSA molar ratio so that with three molar equivalents a saturation was reached, in agreement with the number of high-affinity binding sites reported for the FA. These findings confirm that a non-uniform level of ligand binding in an albumin sample can be an explanation for the early-observed conformational heterogeneity at pH 4.
Acid unfolding; Fatty acid binding; Fluorescence; Human serum albumin; SAXS; Biotechnology; Surfaces and Interfaces; Physical and Theoretical Chemistry; Colloid and Surface Chemistry
01 Pubblicazione su rivista::01a Articolo in rivista
The effect of fatty acid binding in the acid isomerizations of albumin investigated with a continuous acidification method / Del Giudice, Alessandra; Galantini, Luciano; Dicko, Cedric; Pavel, Nicolae V.. - In: COLLOIDS AND SURFACES. B, BIOINTERFACES. - ISSN 0927-7765. - STAMPA. - 168:(2018), pp. 109-116. [10.1016/j.colsurfb.2018.03.038]
File allegati a questo prodotto
File Dimensione Formato  
DelGiudice_The-effect_2018.pdf

solo gestori archivio

Tipologia: Versione editoriale (versione pubblicata con il layout dell'editore)
Licenza: Tutti i diritti riservati (All rights reserved)
Dimensione 1.33 MB
Formato Adobe PDF
1.33 MB Adobe PDF   Visualizza/Apri   Richiedi una copia

I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.

Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11573/1116900
Citazioni
  • ???jsp.display-item.citation.pmc??? 0
  • Scopus 3
  • ???jsp.display-item.citation.isi??? 2
social impact