title: Preferential binding of insecticide phorate with sub-domain IIA of human serum albumin induces protein damage and its toxicological significance.
abstract: Phorate, an organophosphorus insecticide is known for its adverse effects on acetylcholinesterase, and other neuronal and pulmonary activities. Most likely, the toxicity of drugs/agrochemicals is modulated through cellular distribution bound to plasma proteins. Therefore, the in vitro interaction of phorate with human serum albumin (HSA) has been investigated, using sensitive techniques like fluorescence spectroscopy and circular dichroism, to ascertain its binding mechanism and toxicological implications. Fluorescence studies revealed the quenching constant (Ksv) as 2.5 × 10⁴ M⁻¹ and binding affinity (Ka) as 2.96 × 10⁴ M⁻¹ (r² = 0.99), with a primary binding site of phorate at sub-domain IIA of HSA. Circular dichroism (CD) data demonstrated a noticeable reduction in secondary structure (α-helical content) of phorate treated HSA. Albumin treated with 200-1000 μM phorate released significant amounts of acid soluble amino and carbonyl groups, whereas higher concentrations resulted in protein fragmentation. It is postulated that the 1'-O and 3-O alkyl groups of phorate have a role in binding with electrophilic centers of Trp 214, and Arg 218/Lys 195, respectively. Moreover, the significant ultrastructural changes, reactive oxygen species (ROS) generation, mitochondrial damage and cell death in phorate treated cultured human amnion epithelial (WISH) cells, elucidated phorate induced cellular toxicity.
authors.affiliation: A.R. Al-Jeraisy Chair for DNA Research, College of Science, King Saud University, Riyadh 11451, Saudi Arabia.
authors.forename: Quaiser
authors.initials: Q
authors.lastname: Saquib
authors.forename: Abdulaziz A
authors.initials: AA
authors.lastname: Al-Khedhairy
authors.forename: Maqsood A
authors.initials: MA
authors.lastname: Siddiqui
authors.forename: Atanu Singha
authors.initials: AS
authors.lastname: Roy
authors.forename: Swagata
authors.initials: S
authors.lastname: Dasgupta
authors.forename: Javed
authors.initials: J
authors.lastname: Musarrat
mesh_terms: D016923:Cell Death; D002460:Cell Line; D002942:Circular Dichroism / Q000379:methods; D006801:Humans; D008928:Mitochondria / Q000187:drug effects; D008958:Models, Molecular; D018384:Oxidative Stress; D010702:Phorate / Q000737:chemistry* / Q000633:toxicity*; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017382:Reactive Oxygen Species / Q000378:metabolism; D012709:Serum Albumin / Q000737:chemistry*; D013050:Spectrometry, Fluorescence / Q000379:methods; D014364:Tryptophan / Q000737:chemistry
chemical_list: D017382:Reactive Oxygen Species; D012709:Serum Albumin; D010702:Phorate; D014364:Tryptophan