The biological role of human paraoxonase (PON1) remains unclear, whilst there is a consensus that the enzyme has a protective influence. A toxicological role, protecting from environmental poisoning by organophosphate derivatives drove earlier works, and more recently, clinical interest has focused on a protective role in vascular disease. PON1 resides essentially on HDL particles, a complex and dynamic molecular environment. Our recent discovery of the human phosphate binding protein (HPBP), displaying a firm propensity to associate with PON1, has steered new directions for characterizing PON1 functional state. Here, we report investigations on the effect of HPBP on oligomerization, storage and thermal stability of PON1. We found that purified PON1 is as a mixture of at least two states, and that the absence of HPBP favors homo-oligomerization of PON1 into state(s) of higher molecular size. We showed that HPBP allows stabilizing active conformation(s) of PON1 disencumbered of its natural environment. We also showed that PON1 exhibits intrinsically a remarkable thermal stability, and that the association of HPBP strongly contributes to slow the denaturation rate. A hybrid recombinant PON1 was shown more thermostable than the human enzyme, and its stability was unaffected by the presence of HPBP. Altogether, the results strongly encourage further study of the human enzyme. Study holds ProTherm entries: 23060, 23061, 23062, 23063, 23064 Extra Details: Transition 2 (endothermic) Paraoxonase; Protein stability; Catalytic scavenger; Enzyme therapy; Nerve agent; Biopharmaceutical
Submitter: Connie Wang
Submission Date: April 24, 2018, 8:54 p.m.
|Number of data points||5|
|Proteins||Serum paraoxonase/arylesterase 1 ; Serum paraoxonase/arylesterase 1|
|Assays/Quantities/Protocols||Experimental Assay: Tm|
|Libraries||Mutations for sequence MAKLIALTLLGMGLALFRNHQSSYQTRLNALREVQPVELPNCNLVKGIETGSEDLEILPNGLAFISSGLKYPGIKSFNPNSPGKILLMDLNEEDPTVLELGITGSKFDVSSFNPHGISTFTDEDNAMYLLVVNHPDAKSTVELFKFQEEEKSLLHLKTIRHKLLPNLNDIVAVGPEHFYGTNDHYFLDPYLQSWEMYLGLAWSYVVYYSPSEVRVVAEGFDFANGINISPDGKYVYIAELLAHKIHVYEKHANWTLTPLKSLDFNTLVDNISVDPETGDLWVGCHPNGMKIFFYDSENPPASEVLRIQNILTEEPKVTQVYAENGTVLQGSTVASVYKGKLLIGTVFHKALYCEL ; Mutations for sequence MAKLTALTLLGMGLALFDRQKSSFQTRFNVHREVTPVELPNCNLVKGIDNGSEDLEILPNGLAFISSGLKYPGIMSFDPDKSGKILLMDLNEKEPAVSELEIIGNTLDISSFNPHGISTFIDDDNTVYLLVVNHPGSSSTVEVFKFQEEEKSLLHLKTIRHKLLPSVNDIVAVGPEHFYATNDHYFIDPYLKSWEMHLGLAWSFVTYYSPNDVRVVAEGFDFANGINISPDGKYVYIAELLAHKIHVYEKHANWTLTPLRVLSFDTLVDNISVDPVTGDLWVGCHPNGMRIFFYDAENPPGSEVLRIQDILSEEPKVTVVYAENGTVLQGSTVAAVYKGKLLIGTVFHKALYCDL|