Optimisation of myoglobin as a biocatalyst for reversible deactivation radical polymerisation

  • Robert Chadwick

Student thesis: Doctoral Thesis


In this thesis, myoglobin (Mb) is presented as a biocatalyst for reversible-deactivation radical polymerisation (RDRP). Initially, Mb was chemically optimised as a catalyst for atom transfer radical polymerisation (ATRP). The effect of initiator and monomer structure was screened in addition to reaction conditions such as buffer, salt and pH. Mb was found to act as a catalyst for ARGET-ATRP with some degree of control over the polymerisation. Features, such as higher measured number average molar mass (Mn,SEC) than theoretical number average molar mass (Mn,Th) and broad molar mass dispersities (Đ, around 2.0) indicated that the catalytic ability of Mb could still be further optimised.Through biological engineering, including protein and cofactor engineering, a range of Mb mutants and variants were provided and screened under ARGET-ATRP conditions. The Mb variants, with substituted metal cofactors, did not provide enhanced control over the polymerisation. However, several Mb mutants, identified from semi-rational design, showed lower Đ and evidence of increasing Mn,SEC with monomer conversions. The single mutants were further combined to create double and triple mutants, which also showed enhanced control over the ATRP. Mb was also applied as a biocatalyst for the initiation of reversible addition-fragmentation chain-transfer (RAFT) polymerisation. Both the native (pseudo)peroxidase activity and the ability to initiate via an alkyl halide (as in ATRP initiation) were investigated. Mb was able to initiate RAFT polymerisation via both of these mechanisms. Mb mutants, based on more active peroxidase and ATRP activity, were also applied to RAFT polymerisation. Here, the mutants could not initiate via the peroxidase mechanism but were able to initiate via the alkyl halide. Finally, a reversible protein immobilisation strategy was applied in a commercially available glass continuous flow reactor. Both Mb and horseradish peroxidase (HRP) were successfully immobilised in the flow reactor and their peroxidase activity was measured in continuous flow. The immobilisation was proved to be fully reversible through multiple uses of a single reactor. HRP-initiated RAFT initiation was also attempted in continuous flow by decoupling the initiation step to occur in the HRP-immobilised reactor. Unfortunately, this did not yield polymer. Mb is shown to be a versatile and robust biocatalyst for RDRP. By applying molecular biology, Mb can also be tailored to exhibit better performance in these non-native reactions.
Date of Award19 Jan 2022
Original languageEnglish
Awarding Institution
  • University Of Strathclyde
SponsorsUniversity of Strathclyde
SupervisorNico Bruns (Supervisor) & Karen Faulds (Supervisor)

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