title: Organic arsenicals as efficient and highly specific linkers for protein/peptide-polymer conjugation.
abstract: The entropy-driven affinity of trivalent (in)organic arsenicals for closely spaced dithiols has been exploited to develop a novel route to peptide/protein-polymer conjugation. A trivalent arsenous acid (As(III)) derivative (1) obtained from p-arsanilic acid (As(V)) was shown to readily undergo conjugation to the therapeutic peptide salmon calcitonin (sCT) via bridging of the Cys(1)-Cys(7) disulfide, which was verified by RP-HPLC and MALDI-ToF-MS. Conjugation was shown to proceed rapidly (t < 2 min) in situ and stoichiometrically through sequential reduction-conjugation protocols, therefore exhibiting conjugation efficiencies equivalent to those reported for the current leading disulfide-bond targeting strategies. Furthermore, using bovine serum albumin as a model protein, the trivalent organic arsenical 1 was found to demonstrate enhanced specificity for disulfide-bond bridging in the presence of free cysteine residues relative to established maleimide functional reagents. This specificity represents a shift toward potential orthogonality, by clearly distinguishing between the reactivity of mono- and disulfide-derived (vicinal or neighbors-through-space) dithiols. Finally, p-arsanilic acid was transformed into an initiator for aqueous single electron-transfer living radical polymerization, allowing the synthesis of hydrophilic arsenic-functional polymers which were shown to exhibit negligible cytotoxicity relative to a small molecule organic arsenical, and an unfunctionalized polymer control. Poly(poly[ethylene glycol] methyl ether acrylate) (PPEGA480, DPn = 10, Mn,NMR = 4900 g·mol(-1), Đ = 1.07) possessing a pentavalent arsenic acid (As(V)) α-chain end was transformed into trivalent As(III) post-polymerization via initial reduction by biological reducing agent glutathione (GSH), followed by binding of GSH. Conjugation of the resulting As(III)-functional polymer to sCT was realized within 35 min as indicated by RP-HPLC and verified later by thermodynamically driven release of sCT, from the conjugate, in the presence of strong chelating reagent ethanedithiol.
authors: †Department of Chemistry, University of Warwick, Coventry CV4 7AL, United Kingdom.
authors: Paul
authors: P
authors: Wilson
authors: †Department of Chemistry, University of Warwick, Coventry CV4 7AL, United Kingdom.
authors: Athina
authors: A
authors: Anastasaki
authors: †Department of Chemistry, University of Warwick, Coventry CV4 7AL, United Kingdom.
authors: Matthew R
authors: MR
authors: Owen
authors: †Department of Chemistry, University of Warwick, Coventry CV4 7AL, United Kingdom.
authors: Kristian
authors: K
authors: Kempe
authors: †Department of Chemistry, University of Warwick, Coventry CV4 7AL, United Kingdom.
authors: David M
authors: DM
authors: Haddleton
authors: Sarah K
authors: SK
authors: Mann
authors: Angus P R
authors: AP
authors: Johnston
authors: John F
authors: JF
authors: Quinn
authors: †Department of Chemistry, University of Warwick, Coventry CV4 7AL, United Kingdom.
authors: Michael R
authors: MR
authors: Whittaker
authors: §Lowy Cancer Research Centre and Prince of Wales Clinical School, University of New South Wales, Sydney, New South Wales 2052, Australia.
authors: Philip J
authors: PJ
authors: Hogg
authors: †Department of Chemistry, University of Warwick, Coventry CV4 7AL, United Kingdom.
authors: Thomas P
authors: TP
authors: Davis
mesh_terms: D000179:Acrylates / Q000737:chemistry; D000818:Animals; D001152:Arsenicals / Q000138:chemical synthesis / Q000737:chemistry*; D018053:Arsenites / Q000138:chemical synthesis / Q000737:chemistry; D002116:Calcitonin / Q000737:chemistry*; D002460:Cell Line; D003545:Cysteine / Q000737:chemistry*; D051379:Mice; D008958:Models, Molecular; D011092:Polyethylene Glycols / Q000737:chemistry; D058105:Polymerization; D012474:Salmon; D013438:Sulfhydryl Compounds / Q000737:chemistry
chemical_list: D000179:Acrylates; D001152:Arsenicals; D018053:Arsenites; D013438:Sulfhydryl Compounds; D011092:Polyethylene Glycols; C028815:salmon calcitonin; D002116:Calcitonin; C032793:arsenous acid; D003545:Cysteine; C035956:methyl acrylate