RNA-Polymer Hybrids via Direct and Site-Selective Acylation with the ATRP Initiator and Photoinduced Polymerization.

Combining synthetic polymers with RNA paves the way for creating RNA-based materials with non-canonical functions. We have developed an acylation reagent that allows for direct incorporation of the atom transfer radical polymerization (ATRP) initiator into both short synthetic oligoribonucleotides and natural biomass RNA extracted from torula yeast. The acylation was performed in a quantitative yield. The resulting initiator-functionalized RNAs were used for grafting polymer chains from the RNA by photoinduced ATRP, resulting in RNA-polymer hybrids with narrow molecular weight distributions. The RNA initiator was used for the polymerization of oligo(ethylene oxide) methyl ether methacrylate, poly(ethylene glycol) dimethacrylate, and N-isopropylacrylamide monomers, resulting in RNA bottlebrushes, hydrogels, and stimuli-responsive materials. This approach, readily applicable to both post-synthetic and nature-derived RNA, can be used to engineer the properties of a variety of RNA-based macromolecular hybrids and assemblies providing access to a wide variety of RNA-polymer hybrids.

Red-Light-Driven Atom Transfer Radical Polymerization for High-Throughput Polymer Synthesis in Open Air.

Photoinduced reversible-deactivation radical polymerization (photo-RDRP) techniques offer exceptional control over polymerization, providing access to well-defined polymers and hybrid materials with complex architectures. However, most photo-RDRP methods rely on UV/visible light or photoredox catalysts (PCs), which require complex multistep synthesis. Herein, we present the first example of fully oxygen-tolerant red/NIR-light-mediated photoinduced atom transfer radical polymerization (photo-ATRP) in a high-throughput manner under biologically relevant conditions. The method uses commercially available methylene blue (MB+) as the PC and [X-CuII/TPMA]+ (TPMA = tris(2-pyridylmethyl)amine) complex as the deactivator. The mechanistic study revealed that MB+ undergoes a reductive quenching cycle in the presence of the TPMA ligand used in excess. The formed semireduced MB (MB•) sustains polymerization by regenerating the [CuI/TPMA]+ activator and together with [X-CuII/TPMA]+ provides control over the polymerization. This dual catalytic system exhibited excellent oxygen tolerance, enabling polymerizations with high monomer conversions (>90%) in less than 60 min at low volumes (50-250 µL) and high-throughput synthesis of a library of well-defined polymers and DNA-polymer bioconjugates with narrow molecular weight distributions (D < 1.30) in an open-air 96-well plate. In addition, the broad absorption spectrum of MB+ allowed ATRP to be triggered under UV to NIR irradiation (395-730 nm). This opens avenues for the integration of orthogonal photoinduced reactions. Finally, the MB+/Cu catalysis showed good biocompatibility during polymerization in the presence of cells, which expands the potential applications of this method.

Visible-Light-Mediated Controlled Radical Branching Polymerization in Water.

Hyperbranched polymethacrylates were synthesized by green-light-induced atom transfer radical polymerization (ATRP) under biologically relevant conditions in the open air. Sodium 2-bromoacrylate (SBA) was prepared in situ from commercially available 2-bromoacrylic acid and used as a water-soluble inibramer to induce branching during the copolymerization of methacrylate monomers. As a result, well-defined branched polymethacrylates were obtained in less than 30 min with predetermined molecular weights (36 000

Rational Control of Protein-Protein Interactions with Protein-ATRP-Generated Protease-Sensitive Polymer Cages.

Protease-protease interactions lie at the heart of the biological cascades that provide rapid molecular responses to living systems. Blood clotting cascades, apoptosis signaling networks, bacterial infection, and virus trafficking have all evolved to be activated and sustained by protease-protease interactions. Biomimetic strategies designed to target drugs to specific locations have generated proprotein drugs that can be activated by proteolytic cleavage to release native protein. We have previously demonstrated that the modification of enzymes with a custom-designed comb-shaped polymer nanoarmor can shield the enzyme surface and eliminate almost all protein-protein interactions. We now describe the synthesis and characterization of protease-sensitive comb-shaped nanoarmor cages using poly(ethylene glycol) methacrylate macromonomers where the PEG tines of the comb are connected to the backbone of the growing polymer chain by peptide linkers. Protease-induced cleavage of the tines of the comb releases a polymer-modified protein that can once again participate in protein-protein interactions. Atom transfer radical polymerization (ATRP) was used to copolymerize the macromonomer and carboxybetaine methacrylate from initiator-labeled chymotrypsin and trypsin enzymes, yielding proprotease conjugates that retained activity toward small peptide substrates but prevented activity against proteins. Native proteases triggered the release of the PEG side chains from the polymer backbone within 20 min, thereby increasing the activity of the conjugate toward larger protein substrates by 100%. Biomimetic cascade initiation of nanoarmored protease-sensitive protein-polymer conjugates may open the door to a new class of responsive targeted therapies.

Molecular Dynamics-Guided Design of a Functional Protein-ATRP Conjugate That Eliminates Protein-Protein Interactions.

Even the most advanced protein-polymer conjugate therapeutics do not eliminate antibody-protein and receptor-protein recognition. Next-generation bioconjugate drugs will need to replace stochastic selection with rational design to select desirable levels of protein-protein interaction while retaining function. The "Holy Grail" for rational design would be to generate functional enzymes that are fully catalytic with small molecule substrates while eliminating interaction between the protein surface and larger molecules. Using chymotrypsin, an important enzyme that is used to treat pancreatic insufficiency, we have designed a series of molecular chimeras with varied grafting densities and shapes. Guided by molecular dynamic simulations and next-generation molecular chimera characterization with asymmetric flow field-flow fractionation chromatography, we grew linear, branched, and comb-shaped architectures from the surface of the protein by atom-transfer radical polymerization. Comb-shaped polymers, grafted from the surface of chymotrypsin, completely prevented enzyme inhibition with protein inhibitors without sacrificing the ability of the enzyme to catalyze the hydrolysis of a peptide substrate. Asymmetric flow field-flow fractionation coupled with multiangle laser light scattering including dynamic light scattering showed that nanoarmor designed with comb-shaped polymers was particularly compact and spherical. The polymer structure significantly increased protein stability and reduced protein-protein interactions. Atomistic molecular dynamic simulations predicted that a dense nanoarmor with long-armed comb-shaped polymer would act as an almost perfect molecular sieve to filter large ligands from substrates. Surprisingly, a conjugate that was composed of 99% polymer was needed before the elimination of protein-protein interactions.

Catalytic Detoxification of Organophosphorus Nerve Agents by Butyrylcholinesterase-Polymer-Oxime Bioscavengers.

Organophosphorus nerve agents (OPNAs), used in chemical warfare, irreversibly inhibit essential cholinesterases (ChEs) in the cholinergic neurotransmission system. Several potent nucleophilic oximes have been approved for the treatment of acute poisoning by OPNAs, but they are rapidly cleared from blood circulation. Butyrylcholinesterase (BChE) stoichiometrically binds nerve agents, but because the molecular weight of a nerve agent is about 500-fold less than the enzyme, the bioscavenger has had limited utility. We synthesized BChE-polymer-oxime conjugates using atom transfer radical polymerization (ATRP) and azide-alkyne "click" chemistry. The activity of the BChE-polymer-oxime conjugates was dependent on the degree of oxime loading within the copolymer side chains. The covalent modification of oxime-containing copolymers prolonged the activity of BChE in the presence of the VX- and cyclosarin-fluorogenic analogues EMP-MeCyC and CMP-MeCyC, respectively. After complete inactivation by VX and cyclosarin fluorogenic analogues, the conjugates demonstrated efficient self-reactivation of up to 80% within 3-6 h. Repeated inhibition and high-level self-reactivation assays revealed that the BChE-polymer-oxime conjugates were excellent reactivators of OPNA-inhibited BChE. Recurring self-reactivation of BChE-polymer-oxime conjugates following repeated BChE inhibition by fluorogenic OPNAs (Flu-OPNAs) opens the door to developing the next generation of nerve agent "catalytic" bioscavengers.

Charge-Preserving Atom Transfer Radical Polymerization Initiator Rescues the Lost Function of Negatively Charged Protein-Polymer Conjugates.

When grown from the surface of proteins, negatively charged polymers cause irreversible inactivation, thereby limiting the breadth of the synthetic space that negatively charged protein-polymer conjugates can be applied to. More broadly speaking, independent of polymer and synthetic approach, almost all protein-polymer conjugates are less active than their precursors. After more than a decade without major advances in understanding why the attachment of some polymers so sharply deactivates enzymes, we focused our attention on a technique to protect enzymes from the growth of a deactivating polymer by restoring the charge at the protein surface during polymer attachment. We synthesized an amino-reactive positively charged atom transfer radical polymerization initiator that inserted a permanent positive charge at the site of bio-macroinitiator attachment. Preserving the surface charge through attachment of the permanent positively charged initiator led to the first observation of activity of enzymes that were coupled to negatively charged homopolymers.

Molecular Sieving on the Surface of a Nano-Armored Protein.

The molecular sieving properties of protein surface-attached polymers are the central features in how polymers extend therapeutic protein lifetimes in vivo. Yet, even after 30 years of research, permeation rates of molecules through polymer-surrounded protein surfaces are largely unknown. As a result, the generation of protein-polymer conjugates remains a stochastic process, unfacilitated by knowledge of structure-function-polymer architecture relationships. In this work, polymers are grown from the surface of avidin using atom transfer radical polymerization (ATRP) and used to determine how polymer length and density influence the binding kinetics of ligands as a function of ligand size and shape. The rate of binding is strongly dependent on the grafting density of polymers and the size of the ligand but interestingly, far less dependent on the length of the polymer. This study unveils a deeper understanding of relationship between polymer characteristics and binding kinetics, discovering important steps in rational design of protein-polymer conjugates.

Tailoring Site Specificity of Bioconjugation Using Step-Wise Atom-Transfer Radical Polymerization on Proteins.

Protein-polymer conjugates are powerful combinations of the biotic and abiotic worlds that impact many industries. Predicting the site and impact of polymer growth from the surface of proteins is only useful if we can use that information to choose which site to modify synthetically. We have explored the combination of a predictive algorithm with a unique stepwise atom-transfer radical polymerization (ATRP) to selectively move the predominant modification sites around a model enzyme. Lysozyme was modified with defined stoichiometric ratios of polymerization initiators and initiation inhibitors to selectively and strategically grow poly(carboxybetaine methacrylate) polymers from different protein sites. Electrospray ionization mass spectrometry was used to examine the uniformity of the lysozyme-initiator and lysozyme-inhibitor complexes prior to polymer growth. Bioactivity of the lysozyme-polymer conjugates was examined as a function of polymer location on the enzyme surface. Step-wise atom-transfer radical polymerization from proteins provides a versatile and modular approach that can be extended to the rational and selective design of other protein-polymer conjugates.

Intramolecular Interactions of Conjugated Polymers Mimic Molecular Chaperones to Stabilize Protein-Polymer Conjugates.

The power and elegance of protein-polymer conjugates has solved many vexing problems for society. Rational design of these complex covalent hybrids depends on a deep understanding of how polymer physicochemical properties impact the conjugate structure-function-dynamic relationships. We have generated a large family of chymotrypsin-polymer conjugates which differ in polymer length and charge, using grafting-from atom-transfer radical polymerization, to elucidate how the polymers influenced enzyme structure and function at pHs that would unfold and inactivate the enzyme. We also used molecular dynamics simulations to deepen our understanding of protein-polymer intramolecular interactions. Remarkably, the data revealed that, contrary to current thoughts on how polymers stabilize proteins, appropriately designed polymers actually stabilize partially unfolded intermediates and assist in refolding to an active conformation. Long, hydrophilic polymers minimized interfacial interactions in partially unfolded conjugates leading to increased stabilization. The design of covalently attached intramolecular biomimetic chaperones that drive protein refolding could have far reaching consequences.

Effect of Aggressive lipid-lowering treatment with Rosuvastatin on vascular endoTHelium function: evaluation of vascular endothelium function (EARTH study).

Vascular endothelial dysfunction plays an important role in the process of atherosclerosis up to the final stage of plaque rupture. Vascular endothelial dysfunction is reversible, and can be recovered by medications and life-style changes. Improvement in endothelial function may reduce cardiovascular events and improve long-term prognosis. A total of 50 patients with stable angina and dyslipidemia were enrolled, including patients who had not received prior treatment with statins and had serum LDL-C levels ≥ 100 mg/dL, and patients who had previously received statin treatment. All agreed to register regardless of their LDL-C level. Rosuvastatin was initially administered at a dose of 2.5 mg and appropriately titrated up to the maximum dose of 20 mg or until LDL-C levels lower than 80 mg/dL were achieved, for 24 weeks. Endothelial function was assessed by the reactive hyperemia peripheral arterial tonometry (RH-PAT) index in the radial artery by Endo-PAT® 2000 (Endo-PAT®2000, software version 3.0.4, Itamar Medical Ltd., Caesarea, Israel). RH-PAT data were digitally analyzed online by Endo-PAT®2000 at baseline and at 24 weeks. LDL-C and MDA-LDL-C decreased from 112.6 ± 23.3 to 85.5 ± 20.2 mg/dL and from 135.1 ± 36.4 to 113.9 ± 23.5 mg/dL respectively (p < 0.0001). However, HDL-C, hs-CRP and TG did not change significantly after treatment. RH-PAT index levels significantly improved, from 1.60 ± 0.31 to 1.77 ± 0.57 (p = 0.04) after treatment, and the percent change of the RH-PAT index was 12.8 ± 36.9%. Results of multivariate analysis show that serum LDL-C levels over 24 weeks did not act as a predictor of improvement of the RH-PAT index. However, HbA1c at baseline was an independent predictor which influenced the 24-week RH-PAT index level. The RH-PAT index of patients with high HbA1c at baseline did not improve after administration of rosuvastatin but it did improve in patients with low HbA1c at baseline. Aggressive lowering of LDL-C with rosuvastatin significantly improved the RH-PAT index, suggesting that it may improve endothelial function in patients with coronary artery disease.Clinical Trial Registration No: UMIN-CTR, UMIN000010040.

Design of Stomach Acid-Stable and Mucin-Binding Enzyme Polymer Conjugates.

The reduced immunogenicity and increased stability of protein-polymer conjugates has made their use in therapeutic applications particularly attractive. However, the physicochemical interactions between polymer and protein, as well as the effect of this interaction on protein activity and stability, are still not fully understood. In this work, polymer-based protein engineering was used to examine the role of polymer physicochemical properties on the activity and stability of the chymotrypsin-polymer conjugates and their degree of binding to intestinal mucin. Four different chymotrypsin-polymer conjugates, each with the same polymer density, were synthesized using "grafting-from" atom transfer radical polymerization. The influence of polymer charge on chymotrypsin-polymer conjugate mucin binding, bioactivity, and stability in stomach acid was determined. Cationic polymers covalently attached to chymotrypsin showed high mucin binding, while zwitterionic, uncharged, and anionic polymers showed no mucin binding. Cationic polymers also increased chymotrypsin activity from pH 6-8, while zwitterionic polymers had no effect, and uncharged and anionic polymers decreased enzyme activity. Lastly, cationic polymers decreased the tendency of chymotrypsin to structurally unfold at extremely low pH, while uncharged and anionic polymers induced unfolding more quickly. We hypothesized that when polymers are covalently attached to the surface of a protein, the degree to which those polymers interact with the protein surface is the predominant determinant of whether the polymer will stabilize or inactivate the protein. Preferential interactions between the polymer and the protein lead to removal of water from the surface of the protein, and this, we believe, inactivates the enzyme.

Bactericidal Specificity and Resistance Profile of Poly(Quaternary Ammonium) Polymers and Protein-Poly(Quaternary Ammonium) Conjugates.

Antibacterial polymers are potentially powerful biocides that can destroy bacteria on contact. Debate in the literature has surrounded the mechanism of action of polymeric biocides and the propensity for bacteria to develop resistance to them. There has been particular interest in whether surfaces with covalently coupled polymeric biocides have the same mechanism of action and resistance profile as similar soluble polymeric biocides. We designed and synthesized a series of poly(quaternary ammonium) polymers, with tailorable molecular structures and architectures, to engineer their antibacterial specificity and their ability to delay the development of bacterial resistance. These linear poly(quaternary ammonium) homopolymers and block copolymers, generated using atom transfer radical polymerization, had structure-dependent antibacterial specificity toward Gram positive and negative bacterial species. When single block copolymers contained two polymer segments of differing antibacterial specificity, the polymer combined the specificities of its two components. Nanoparticulate human serum albumin-poly(quaternary ammonium) conjugates of these same polymers, synthesized via "grafting from" atom transfer radical polymerization, were strongly biocidal and also exhibited a marked decrease in the rate of bacterial resistance development relative to linear polymers. These protein-biocide conjugates mimicked the behavior of surface-presented polycationic biocides rather than their nonproteinaceous counterparts.

Dramatically increased pH and temperature stability of chymotrypsin using dual block polymer-based protein engineering.

In this study, we report on multimodal temperature-responsive chymotrypsin-poly(sulfobetaine methacrylamide)-block-poly(N-isopropylacrylamide) (CT-pSBAm-block-pNIPAm) protein-polymer conjugates. Using polymer-based protein engineering (PBPE) with aqueous atom transfer radical polymerization (ATRP), we synthesized three different molecular weight CT-pSBAm-block-pNIPAm bioconjugates that responded structurally to both low and high temperature. In the block copolymer grown from the surface of the enzyme, upper critical solution temperature (UCST) phase transition was dependent on the chain length of the polymers in the conjugates, whereas lower critical solution temperature (LCST) phase transition was independent of molecular weight. Each CT-pSBAm-block-pNIPAm conjugate showed temperature dependent changes in substrate affinity and productivity when assayed from 0 to 40 °C. In addition, these conjugates showed higher stability to harsh conditions, including temperature, low pH, and protease degradation. Indeed, the PBPE-modified enzyme was active for over 8 h in the presence of a stomach protease at pH 1.0. Using PBPE, we created a dual zone shell surrounding each molecule of enzyme. The thickness of each zone of the shell was engineered to be separately responsive to temperature.

Rational tailoring of substrate and inhibitor affinity via ATRP polymer-based protein engineering.

Atom transfer radical polymerization (ATRP)-based protein engineering of chymotrypsin with a cationic polymer was used to tune the substrate specificity and inhibitor binding. Poly(quaternary ammonium) was grown from the surface of the enzyme using ATRP after covalent attachment of a protein reactive, water-soluble ATRP-initiator. This "grafting from" conjugation approach generated a high density of cationic ammonium ions around the biocatalytic core. Modification increased the surface area of the protein over 40-fold, and the density of modification on the protein surface was approximately one chain per 4 nm(2). After modification, bioactivity was increased at low pH relative to the activity of the native enzyme. In addition, the affinity of the enzyme for a peptide substrate was increased over a wide pH range. The massively cationic chymotrypsin, which included up to 2000 additional positive charges per molecule of enzyme, was also more stable at extremes of temperature and pH. Most interestingly, we were able to rationally control the binding of two oppositely charged polypeptide protease inhibitors, aprotinin and the Bowman-Birk trypsin-chymotrypsin inhibitor from Glycine max, to the cationic derivative of chymotrypsin. This study expands upon our efforts to use polymer-based protein engineering to predictably engineer enzyme properties without the need for molecular biology.

Tailored Branched Polymer-Protein Bioconjugates for Tunable Sieving Performance.

Protein-polymer conjugates combine the unique properties of both proteins and synthetic polymers, making them important materials for biomedical applications. In this work, we synthesized and characterized protein-branched polymer bioconjugates that were precisely designed to retain protein functionality while preventing unwanted interactions. Using chymotrypsin as a model protein, we employed a controlled radical branching polymerization (CRBP) technique utilizing a water-soluble inibramer, sodium 2-bromoacrylate. The green-light-induced atom transfer radical polymerization (ATRP) enabled the grafting of branched polymers directly from the protein surface in the open air. The resulting bioconjugates exhibited a predetermined molecular weight, well-defined architecture, and high branching density. Conformational analysis by SEC-MALS validated the controlled grafting of branched polymers. Furthermore, enzymatic assays revealed that densely grafted polymers prevented protein inhibitor penetration, and the resulting conjugates retained up to 90% of their enzymatic activity. This study demonstrates a promising strategy for designing protein-polymer bioconjugates with tunable sieving behavior, opening avenues for applications in drug delivery and biotechnology.

Bottlebrush Polymers for Articular Joint Lubrication: Influence of Anchoring Group Chemistry on Lubrication Properties.

The role of carboxylic, aldehyde, or epoxide groups incorporated into bottlebrush macromolecules as anchoring blocks (or cartilage-binding blocks) is investigated by measuring their lubricating properties and cartilage-binding effectiveness. Mica modified with amine groups is used to mimic the cartilage surface, while bottlebrush polymers functionalized with carboxylic, aldehyde, or epoxide groups played the role of the lubricant interacting with the cartilage surface. We demonstrate that bottlebrushes with anchoring blocks effectively reduce the friction coefficient on modified surfaces by 75-95% compared to unmodified mica. The most efficient polymer appears to be the one with epoxide groups, which can react spontaneously with amines at room temperature. In this case, the value of the friction coefficient is the lowest and equals 0.009 ± 0.001, representing a 95% reduction compared to measurements on nonmodified mica. These results show that the presence of the functional groups within the anchoring blocks has a significant influence on interactions between the bottlebrush polymer and cartilage surface. All synthesized bottlebrush polymers are also used in the preliminary lubrication tests carried out on animal cartilage surfaces. The developed materials are very promising for future in vivo studies to be used in osteoarthritis treatment.

Hydrophilic Poly(meth)acrylates by Controlled Radical Branching Polymerization: Hyperbranching and Fragmentation.

Topology significantly impacts polymer properties and applications. Hyperbranched polymers (HBPs) synthesized via atom transfer radical polymerization (ATRP) using inimers typically exhibit broad molecular weight distributions and limited control over branching. Alternatively, copolymerization of inibramers (IB), such as α-chloro/bromo acrylates with vinyl monomers, yields HBPs with precise and uniform branching. Herein, we described the synthesis of hydrophilic HB polyacrylates in water by copolymerizing a water-soluble IB, oligo(ethylene oxide) methyl ether 2-bromoacrylate (OEOBA), with various hydrophilic acrylate comonomers. Visible-light-mediated controlled radical branching polymerization (CRBP) with dual catalysis using eosin Y (EY) and copper complexes resulted in HBPs with various molecular weights (M n = 38 000 to 170 000) and degrees of branching (2%-24%). Furthermore, the optimized conditions enabled the successful application of the OEOBA to synthesize linear-hyperbranched block copolymers and hyperbranched polymer protein hybrids (HB-PPH), demonstrating its potential to advance the synthesis of complex macromolecular architecture under environmentally benign conditions. Copolymerization of hydrophilic methacrylate monomer, oligo(ethylene oxide) methyl ether methacrylate (OEOMA500), and inibramer OEOBA was accompanied by fragmentation via ß-carbon C-C bond scission and subsequent growth of polymer chains from the fragments. Furthermore, computational studies investigating the fragmentation depending on the IB and comonomer structure supported the experimental observations. This work expands the toolkit of water-soluble inibramers for CRBP and highlights the critical influence of the inibramer structure on reaction outcomes.

Split fluorescent protein-mediated multimerization of cell wall binding domain for highly sensitive and selective bacterial detection.

Cell wall peptidoglycan binding domains (CBDs) of cell lytic enzymes, including bacteriocins, autolysins and bacteriophage endolysins, enable highly selective bacterial binding, and thus, have potential as biorecognition molecules for nondestructive bacterial detection. Here, a novel design for a self-complementing split fluorescent protein (FP) complex is proposed, where a multimeric FP chain fused with specific CBDs ((FP-CBD)n) is assembled inside the cell, to improve sensitivity by enhancing the signal generated upon Staphylococcus aureus or Bacillus anthracis binding. Flow cytometry shows enhanced fluorescence on the cell surface with increasing FP stoichiometry and surface plasmon resonance reveals nanomolar binding affinity to isolated peptidoglycan. The breadth of function of these complexes is demonstrated through the use of CBD modularity and the ability to attach enzymatic detection modalities. Horseradish peroxidase-coupled (FP-CBD)n complexes generate a catalytic amplification, with the degree of amplification increasing as a function of FP length, reaching a limit of detection (LOD) of 103 cells/droplet (approximately 0.1 ng S. aureus or B. anthracis) within 15 min on a polystyrene surface. These fusion proteins can be multiplexed for simultaneous detection. Multimeric split FP-CBD fusions enable use as a biorecognition molecule with enhanced signal for use in bacterial biosensing platforms.

Aqueous photo-RAFT polymerization under ambient conditions: synthesis of protein-polymer hybrids in open air.

A photoinduced reversible addition-fragmentation chain-transfer (photo-RAFT) polymerization technique in the presence of sodium pyruvate (SP) and pyruvic acid derivatives was developed. Depending on the wavelength of light used, SP acted as a biocompatible photoinitiator or promoter for polymerization, allowing rapid open-to-air polymerization in aqueous media. Under UV irradiation (370 nm), SP decomposes to generate CO2 and radicals, initiating polymerization. Under blue (450 nm) or green (525 nm) irradiation, SP enhances the polymerization rate via interaction with the excited state RAFT agent. This method enabled the polymerization of a range of hydrophilic monomers in reaction volumes up to 250 mL, eliminating the need to remove radical inhibitors from the monomers. In addition, photo-RAFT polymerization using SP allowed for the facile synthesis of protein-polymer hybrids in short reaction times (<1 h), low organic content (≤16%), and without rigorous deoxygenation and the use of transition metal photocatalysts. Enzymatic studies of a model protein (chymotrypsin) showed that despite a significant loss of protein activity after conjugation with RAFT chain transfer agents, the grafting polymers from proteins resulted in a 3-4-fold recovery of protein activity.