Polymer-Based Protein Engineering: Synthesis and Characterization of Armored, High Graft Density Polymer-Protein Conjugates

Sheiliza Carmali, Hironobu Murata, Chad Cummings, Krzysztof Matyjaszewski, Alan J Russell

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

Atom transfer radical polymerization (ATRP) from the surface of a protein can generate remarkably dense polymer shells that serve as armor and rationally tune protein function. Using straightforward chemistry, it is possible to covalently couple or display multiple small molecule initiators onto a protein surface. The chemistry is fine-tuned to be sequence specific (if one desires a single targeted site) at controlled density. Once the initiator is anchored on the protein surface, ATRP is used to grow polymers on protein surface, in situ. The technique is so powerful that a single-protein polymer conjugate molecule can contain more than 90% polymer coating by weight. If desired, stimuli-responsive polymers can be "grown" from the initiated sites to prepare enzyme conjugates that respond to external triggers such as temperature or pH, while still maintaining enzyme activity and stability. Herein, we focus mainly on the synthesis of chymotrypsin-polymer conjugates. Control of the number of covalently coupled initiator sites by changing the stoichiometric ratio between enzyme and the initiator during the synthesis of protein-initiator complexes allowed fine-tuning of the grafting density. For example, very high grafting density chymotrypsin conjugates were prepared from protein-initiator complexes to grow the temperature-responsive polymers, poly(N-isopropylacrylamide), and poly[N,N'-dimethyl(methacryloyloxyethyl) ammonium propane sulfonate]. Controlled growth of polymers from protein surfaces enables one to predictably manipulate enzyme kinetics and stability without the need for molecular biology-dependent mutagenesis.

Original languageEnglish
Pages (from-to)347-380
JournalMethods in Enzymology
Volume590
DOIs
Publication statusPublished - 16 Feb 2017
Externally publishedYes

Bibliographical note

© 2017 Elsevier Inc. All rights reserved.

Keywords

  • Acrylamides/chemistry
  • Acrylic Resins/chemistry
  • Biocatalysis
  • Chymotrypsin/chemistry
  • Enzyme Stability
  • Enzymes, Immobilized/chemistry
  • Kinetics
  • Polymerization
  • Proteolysis

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