Sortase-Mediated Surface Functionalization of Stimuli-Responsive Microgels

Abstract

In this work we explored an enzyme-mediated method for selective and efficient decoration of aqueous microgels with biomolecules. Poly(N-vinylcaprolactam) (VCL) microgels with varied amounts of glycidyl methacrylate (GMA) as comonomer incorporated in the microgel shell were synthesized and characterized in regard to their size, swelling degree, and temperature-responsiveness in aqueous solutions. The surface of the PVCL/GMA microgel containing 5 mol % glycidyl methyacrylate was modified by grafting of a specific recognition peptide sequence (LPETG) for Sortase A from Staphylococcus aureus (Sa-SrtA). Sortase-mediated conjugation of the enhanced Green Fluorescent Protein (eGFP) carrying a N-terminal triglycine tag to LPETG-modified microgels was successfully performed. Conjugation of eGFP to the microgel surface was qualitatively proven by confocal microscopy and by fluorescence intensity measurements. The developed protocol enables a precise control of the amount of eGFP grafted to the microgel surface as evidenced by the linear increase of fluorescence intensity of modified microgel samples. The kinetic of the sortase-mediated coupling reaction was determined by time-dependent fluorescence intensity measurements. In summary, sortase-mediated coupling reactions are a simple and powerful technique for targeted surface functionalization of stimuli-responsive microgels with biomolecules.

Synthesis of Polyampholyte Janus‐like Microgels by Coacervation of Reactive Precursors in Precipitation Polymerization

“…[25] It was also used here to study the morphology of the Janus-like microgels in aqueous solution. Before measuring, Janus-like microgels were labeled with a negatively charged fluorescent dye sodium 5,5'-((perfluorocyclopent-1-ene-1,2-diyl)bis(2-ethyl-1,1-dioxidobenzo [b]thiophene-3,6-diyl))bis(2-methoxybenzenesulfonate) [26] that effectively attaches to the imidazole group at pH 4 (Figure 3 B). Figure 3 A reveals the morphology of Janus-like microgels under acidic condition (pH 4), where protonation of imidazole groups take place, leading to a more prominent swelling of the NIPAm-VIm based side.…”

Amylose‐Coated Biohybrid Microgels by Phosphorylase‐Catalyzed Grafting‐From Polymerization

“…In our group, recently a method for selective surface‐modification of microgels with biomacromolecules was developed, the so called sortase‐mediated ligation (SML). After having covalently attached the peptide sequence LPETG for recognition by the transpeptidase enzyme sortase A (SrtA) to PVCL microgels, SrtA is added to enzymatically attach an oligoglycin‐tagged protein or enzyme . Besides, also the conjugation of DNA oligonucleotides to microgels was already reported.…”

“…After having covalently attached the peptide sequence LPETG for recognition by the transpeptidase enzyme sortase A (SrtA) to PVCL microgels, SrtA is added to enzymatically attach an oligoglycin-tagged protein or enzyme. [14] Besides, also the conjugation of DNA oligonucleotides to microgels was already reported. Ali et al described the covalent coupling of 5′-amine modified DNA oligonucleotides to carboxyl-containing PNIPAm microgels via EDC/N-hydroxysulfosuccinimide (NHSS) chemistry.…”

In Situ Growth and Size Regulation of Single Gold Nanoparticles in Composite Microgels

“…The big advantage of PVCL in comparison to PNIPAAm is that it shows superior biocompatibility . Numerous other monomers and cross‐linkers are known for their integration in the polymer network, to define the chemical composition of the MGs and to introduce various functional groups . Thus, beside the temperature‐induced volume change, MGs are able to swell or shrink when triggered by other external stimuli like pH, ionic strength, or light .…”

“…Consequently, a multitude of possible applications in biotechnology, biomedicine (imaging agents, drug carriers, etc. ), and biocatalysis are conceivable …”