Human platelet acetylcholinesterase inhibition by cyclophosphamide: a combined experimental and computational approach.
Acetylcholinesterase (AChE)-inhibition is an area of priority research as various roles have been attributed to AChE in neurodegenerative disorders and cancer as well. In the present study, a comparative multiple 4 dimensional (4D)-approach was applied to analyze human platelet AChE-inhibition by cyclophosphamide (CP). AChE activity was assessed by measuring the hydrolysis of acetylthiocholine iodide (ASChI). The different perspective of analysis was based on two classical (Lineweaver-Burk as well as Dixon) plots, built-in equations of GOSA and a recently introduced graphical approach. Thus, various kinetic constants such as KI, Ks, Km, ksl, Vmao, Ki, ksli, Slmax, �Ks, K1/2, kcat and ksp were estimated. Previous findings of AChE (from different sources) inhibition by CP were also compared. This study extends the elucidation of the kinetic approach of analysis and quantifying enzyme-substrate and enzyme-inhibitor interactions, which is crucial to bringing any drug from bench to bedside. The acyl pocket of human AChE was found to interact with CP through the amino acid residues Y70, Y121, W233, F288, F290, Y334, F408 and Y442, while the anionic sub-site of catalytic site (CAS) interacted with the ligand through residues W84, N85, G116, G117, Y121, S122, G123, L127, Y130, E198, Y334, H443 and G444. CP displayed variable docking poses with the peripheral anionic site (PAS) of human AChE. The findings of kinetic analysis were reinforced by the results of docking experiments. Both the applied approaches strongly indicate partial mixed type of inhibition pattern for the study enzyme (AChE) and its inhibitor (CP).
King Fahd Medical Research center, King Abdulaziz University, Jeddah, Saudi Arabia.
Abdulaziz A
AA
Al-Jafari
Shazi
S
Shakil
Marcella
M
Reale
Mohammad A
MA
Kamal
D000110:Acetylcholinesterase / Q000378:metabolism*; D000328:Adult; D001792:Blood Platelets / Q000187:drug effects* / Q000201:enzymology*; D002800:Cholinesterase Inhibitors / Q000494:pharmacology*; D019295:Computational Biology / Q000379:methods*; D003520:Cyclophosphamide / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D006801:Humans; D008297:Male; D013379:Substrate Specificity / Q000187:drug effects / Q000502:physiology
D002800:Cholinesterase Inhibitors; D003520:Cyclophosphamide; D000110:Acetylcholinesterase