Smart PEGylation of Trypsin

Abstract

Thermoresponsive oligo(ethylene glycol)-based copolymers were investigated for trypsin conjugation. These copolymers have been synthesized by atom transfer radical polymerization of 2-(2-methoxyethoxy)ethyl methacrylate (MEO(2)MA) with oligo(ethylene glycol) methyl ether methacrylate (OEGMA(475), M(n) = 475 g.mol(-1)) at 60 degrees C in the presence of copper(I) chloride and 2,2'-bipyridyl. Two different ATRP initiators, containing succinimidyl ester moieties, were tested, namely, N-succinimidyl-2-bromopropionate and N-succinimidyl-2-bromoisobutyrate. In both cases, ATRP afforded well-defined polymers with a narrow molecular weight distribution and controlled chain-ends. However, the efficiency of initiation of the two initiators was lower than 1 and therefore the formed polymers exhibited a higher than expected mean degree of polymerization. Nevertheless, all types of polymers could be conjugated to trypsin. The conjugation reaction was performed in borax-HCl buffer. Sodium dodecyl sulfate poly(acrylamide) gel electrophoresis (SDS-PAGE) indicated that polymer/enzyme conjugates were obtained in all cases. However, (co)polymers initiated by N-succinimidyl-2-bromopropionate led to the best conjugation results. The formed P(MEO(2)MA-co-OEGMA(475))-trypsin conjugates were found to be thermoresponsive and moreover exhibited a higher enzymatic activity than unmodified trypsin.

Thermo‐Switchable Materials Prepared Using the OEGMA‐Platform

“…We reported lately the modifi cation of bovine pancreas trypsin with thermoresponsive oligo(ethylene glycol)-based (co)polymers (Figure 1 ). [ 49 ] In this approach, thermoresponsive P(MEO 2 MA-co -OEGMA 475 ) copolymers containing N -hydroxysuccinimide active ester α -chain-ends were reacted with the surface lysine residues of trypsin (i.e., grafting-onto approach). The formed P(MEO 2 MA-co -OEGMA 475 )-trypsin conjugates were found to be thermoresponsive and moreover exhibited a higher enzymatic activity than unmodifi ed trypsin.…”

Conformation–function relationships for the comb-shaped polymer pOEGMA

“…In addition to providing a steric barrier to protein adsorption, the hydrophobicity of the pOEGMA main-chain has been found in some cases to enhance activity, possibly by creating a hydrophobic microenvironment that promotes the sequestration of hydrophobic substrates. This was notably observed for a bio-conjugate of P2-stat-P9 and trypsin [74]. pOEGMA has also been used for similar purposes to protect DNA (or RNA) within self-assembled supramolecular structures such as complex coacervate core micelles or by covalent conjugation [75][76][77].…”

Synthesis of novel temperature responsive PEG-b-[PCL-g-P(MEO2MA-co-OEGMA)]-b-PEG (tBG) triblock-graft copolymers and preparation of tBG/graphene oxide composite hydrogels via click chemistry

“…Novel 37 37 a n mPEG :n αClεCL :n εCL is the initial molar ratio of mPEG, αClεCL and εCL. b GD is the grafting density of tBG polymers, which is determined by 1 H NMR of tBG.…”

“…Since P(MEO 2 MA-co-OEGMA) is a hydrophilic temperature responsive polymer in nature [18,37], after the P(MEO 2 MA -co-OEGMA) was grafted on the hydrophobic PCL block of PEG-b-PCL-b-PEG, water-solubility of the synthesized triblock-graft copolymers (tBGs) should be improved and endowed with temperature sensitivity. The density of P(MEO 2 MA-co-OEGMA) graft chains has been adjusted to study its effects on water solubility of the prepared tBGs.…”