1. Synthesis and Characterization of Porphyrin Nanoparticles to model Heme Protein Iron Coordination
Graham Beaton , Samuel Pazicni Parsons Hall, 23 Academic Way, Durham NH 03824
Monomer Characterization
Introduction
Nanoparticle Collapse
Heme proteins serve many vital functions in biological systems, including oxygen delivery, electron transport and redox catalysis. Their function depends largely on the reactivity of the iron metal center, which is not only influenced by covalent interactions of the Heme prosthetic group but also by interactions of the secondary coordination sphere. A Porphyrin cored nanoparticle was synthesized to model heme cofactor iron coordination and the effects of the Heme’s secondary coordination sphere on the iron center’s reactivity were explored. Several methacrylate monomers were synthesized and subjected to RAFT polymerization with porphyrin cored CTA to create a functionalized porphyrin cored polymer. An Iron insertion incorporated the metal center into the porphyrin cored polymer and upon photodimerization with 350 nm light, it was collapsed into a nanoparticle where functionalized chains allowed for a tunable microenvironment around the metal center.
Figure 1: 400 MHz 1H NMR of AMMA
Figure 2: 400 MHz 1H NMR of PFPMA = = =
H2O
Figure 6: Photodimerization of AMMA units with 350 nm light facilitating nanoparticle collapse
Nanoparticle collapse was monitored by UV-Vis (5 min. increments) and Gel Permeation Chromatography (10 min. increments) Hypochromic shift in Anthracene bands and increased retention time are indicative of successful collapse
Polymer Synthesis and Characterization
Monomer Synthesis
Figure 7: UV-Vis of PCNP collapse
Figure 8: GPC of PCNP collapse
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Fe(III) center was reduced to Fe(III) with Dithionite and binding studies were run with Imidazole and Carbon monoxide as axial bound ligands. This binding was monitored with UV-Vis and the soret shifts were consistent with reduced and small molecule bound species.
Monomers were chosen based on their susceptibility to RAFT polymerization, ability to facilitate nanoparticle collapsing and functionality to afford realistic protein microenvironments.
Acrylamide System
Incorporation of PFPMA and AMMA units allow for photodimerization collapse mechanism as well as post polymerization modifications via amine activated ester functionality
Figure 9: UV-Vis spectroscopy of reduced and ligated PCNP species
(1) C
Future Work
With the methacrylate polymer system made and conditions understood further work can be done to probe the reactivity of the metal center. The activated ester functionality of the PFPMA units afford a polymer susceptible to many post polymerization modifications, this will allow the secondary coordination sphere to be tuned towards hydrophobic, hydrophilic or hydrogen bond rich microenvironments. This will also be accomplished with further development of the acrylamide based polymer system. B B
(2) A C A
Methacrylate System
Polymer Backbone (B)
Figure 1: 400 MHz 1H NMR of Por (PFPMA Co AMMA) at 12% AMMA incorporation
Iron Insertion
Acknowledgements
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Deprotonation of the pyrrole Nitrogens with 2,4,6-colidine afford the free base porphyrin core, upon metalation with Iron Bromide, the iron cored star polymer is synthesized and the reaction is monitored via UV-Vis spectroscopy.
Dr. Erik Berda Dr. Kyle J. Rodriguez Dr. Roy Planalp
Hamel Center SURF Fellowship
UNH Instrumentation Center Dr. Patricia Wilkinson
Metalated star polymer
References
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Free-base star polymer
Figure 3: UV-Vis spectrum of PCSP pre and post iron insertion
Acrylamide monomers (1),(2) Purified via Liquid-Liquid extraction, Organic phase washed with H2O and Brine, dried over MgSO4 and isolated via column chromatography 2:1 Hexanes , Ethyl Acetate
(3) Organic phase washed with H2O, Brine, Sodium Bicarbonate and 1M HC
(4) Vaccum filtered to remove DCU, isolated via column chromatography 5:1 Toluene , Ethyl Acetate
Hyperchromic shift of alpha, Beta Q bands and the hypochromic shift of soret band are consistent with metalation of porphyrin with Fe(II)