Interaction of ochratoxin A with human serum albumin. Binding sites localized by competitive interactions with the native protein and its recombinant fragments.
Competitive interactions of ochratoxin A (OTA) and several other acidic compounds were utilized to gain insight into the localization of binding sites and the nature of binding interactions between anionic species and human serum albumin (HSA). Depolarization of OTA fluorescence in the presence of a competing anion was used to quantify ligand-protein interactions. The results obtained were rationalized in terms of OTA displacement from its major binding site. Based on their ability to displace OTA, two distinct groups of the anionic ligands were revealed. The first group contained structurally diverse compounds that shared a common binding site in subdomain IIA (Sudlow Site I). The second group consisted of three non-steroidal anti-inflammatory drugs, which showed much lower affinity to Site I than the OTA dianion. The major site for these drugs was located in domain III. Fluorescence spectroscopy measurements of OTA, warfarin (WAR) and naproxen (NAP) complexes with recombinant proteins corresponding to the domains of HSA (D1-D3) revealed binding to all domains but with different affinities. The binding constants for OTA and WAR decreased in the series D2z.Gt;D3>D1. In contrast, NAP showed the most favorable interaction with D3 and comparable affinities to the two remaining domains. The OTA binding constant for D2, 7.9 x 10(5) M(-1), was smaller than the largest constant for HSA by a factor of approximately 7. The binding constant for OTA with D3, 1.1 x 10(5) M(-1), was very close to that of the secondary binding site for HSA.
Il'ichev
Yuri V
YV
Department of Chemistry, Duke University, Durham, NC 27708, USA.
Perry
Jennifer L
JL
Rüker
Florian
F
Dockal
Michael
M
Simon
John D
JD
D000838:Anions / Q000378:metabolism; D001667:Binding, Competitive; D005454:Fluorescence Polarization; D006801:Humans; D008024:Ligands; D008958:Models, Molecular; D015394:Molecular Structure; D009288:Naproxen / Q000378:metabolism; D009793:Ochratoxins / Q000737:chemistry / Q000378:metabolism*; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012709:Serum Albumin / Q000737:chemistry / Q000378:metabolism*; D014859:Warfarin / Q000378:metabolism
D000838:Anions; D008024:Ligands; D009793:Ochratoxins; D010446:Peptide Fragments; D011994:Recombinant Proteins; D012709:Serum Albumin; C025589:ochratoxin A; D009288:Naproxen; D014859:Warfarin