1. Polymeric dibromomaleimides as extremely efficient disulfide bridging bioconjugation and pegylation agents
Mathew W Jones, Rachel A Strickland, Felix F Schumacher, Stephen Caddick, James R Baker, Matthew I Gibson, David M Haddleton J Am Chem Soc. 2012 Jan 25;134(3):1847-52. doi: 10.1021/ja210335f. Epub 2012 Jan 13.
A series of dibromomaleimides have been shown to be very efficacious at insertion into peptidic disulfide bonds. This conjugation proceeds with a stoichiometric balance of reagents in buffered solutions in less than 15 min to give discrete products while maintaining the disulfide bridge and thus peptide conformation. The insertion is initiated by disulfide reduction using a water-soluble phosphine, tris(2-carboxyethyl)phosphine (TCEP) which allows for subsequent substitution of the two maleimide bromides by the generated thiols. Reaction of salmon calcitonin (sCT) with 2,3-dibromomaleimide (1.1 excess) in the presence of TCEP (1.1 equiv) in aqueous solution at pH 6.2 gives complete production of a single conjugate which requires no workup. A linear methoxy poly(ethylene glycol) (PEG) was functionalized via a Mitsunobu reaction and used for the successful site-specific and rapid pegylation of sCT. This reaction occurs in 15 min with a small stoichiometry excess of the pegylating agent to give insertion at the disulfide with HPLC showing a single product and MALDI-ToF confirming conjugation. Attempts to use the group in a functional ATRP polymerization initiator led to polymerization inhibition. Thus, in order to prepare a range of functional polymers an indirect route was chosen via both azide and aniline functional initiators which were converted to 2,3-dibromomaleimides via appropriate reactions. For example, the azide functional polymer was reacted via a Huisgen CuAAC click reaction to an alkyne functional 2,3-dibromomaleimide. This new reagent allowed for the synthesis of conjugates of sCT with comb polymers derived from PEG methacrylic monomers which in addition gave appropriate cloud points. This reaction represents a highly efficient polymer conjugation method which circumvents problems of purification which normally arise from having to use large excesses of the conjugate. In addition, the tertiary structure of the peptide is efficiently maintained.
2. Thio-Bromo "Click" Reaction Derived Polymer-Peptide Conjugates for Their Self-Assembled Fibrillar Nanostructures
Sonu Kumar, Gerd Hause, Wolfgang H Binder Macromol Biosci. 2020 Jun;20(6):e2000048. doi: 10.1002/mabi.202000048. Epub 2020 Apr 13.
The synthesis and self-assembly of peptide-polymer conjugates into fibrillar nanostructures are reported, based on the amyloidogenic peptide KLVFF. A strategy for rational synthesis of polymer-peptide conjugates is documented via tethering of the amyloidogenic peptide segment LVFF (Aβ17-20 ) and its modified derivative FFFF to the hydrophilic poly(ethylene glycol) monomethyl ether (mPEG) polymer via thio-bromo based "click" chemistry. The resultant conjugates mPEG-LVFF-OMe and mPEG-FFFF-OMe are purified via preparative gel permeation chromatography technique (with a yield of 61% and 64%, respectively), and are successfully characterized via combination of spectroscopic and chromatographic methods, including electrospray ionization time-of-flight mass spectrometry. The peptide-guided self-assembling behavior of the as-constructed amphiphilic supramolecular materials is further investigated via transmission electron microscopic and circular dichroism spectroscopic analysis, exhibiting fibrillar nanostructure formation in binary aqueous solution mixture.
3. One-Pot Synthesis of Thermoresponsive Amyloidogenic Peptide-Polymer Conjugates via Thio-Bromo "Click" Reaction of RAFT Polymers
Sonu Kumar, Stefanie Deike, Wolfgang H Binder Macromol Rapid Commun. 2018 Jan;39(2). doi: 10.1002/marc.201700507. Epub 2017 Oct 27.
A synthetic strategy to efficiently prepare main-chain peptide-polymer conjugates probing their aggregation in solution is described. An in situ tandem reaction based on aminolysis/thio-bromo "click" reaction is performed to tether an amyloidogenic peptide fragment amyloid-β17-20 (Leu-Val-Phe-Phe (LVFF)) to the ω-chain end of poly(diethylene glycol methyl ether acrylate) (PDEGA), prepared via reversible addition fragmentation chain transfer polymerization. Structural confirmation of the constructed conjugates PDEGA-LVFF (Mn,SEC = 5600, Ð = 1.21), (Mn,SEC = 7600, Ð = 1.16), and (Mn,SEC = 8900, Ð = 1.15) is successfully made by combined studies of 1 H NMR, size-exclusion chromatography, matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry, and electrospray ionization time-of-flight (ESI-TOF) mass spectrometry. The effect of the peptidic constituent on the thermoresponsive behavior of the polymer is examined by UV-vis spectroscopy, and the self-assembly behavior of the amphiphilic conjugate is further exploited, exhibiting micellar morphology in aqueous solution.
