Molecular Sieving with PEGylated Dendron-Protein Conjugates

A series of generation 3-5 dendrons based on abis(2,2-hydroxymethylpropionicacid) (bis-MPA) scaffold bearing three respective lengths of linearpoly(ethylene glycol) at their periphery and a dibenzocyclooctyneunit at their core was prepared. These dendrons were appended to thesurface of azide-decorated & alpha;-chymotrypsin (& alpha;-CT) via strain-promotedazide-alkyne cycloaddition to yield a library of dendron-proteinconjugates. These conjugates were characterized by FT-IR and NMR spectroscopyand were imaged using cryo-electron microscopy. The activity of thePEGylated & alpha;-CT-dendron conjugates was investigated using a smallmolecule (benzoyl-l-tyrosine p-nitroanilide)as well as different proteins of different sizes and crystallinities(casein and bovine serum albumin) as substrates. It was found thatthe activity of the conjugates toward the small molecule was largelyretained, while the activity toward the proteins was significantlydiminished. Furthermore, the results indicate that for most of theconjugates the PEG length had a more pronounced impact on enzyme activitythan the dendron generation. Overall, the highest sieving ratios werefound for & alpha;-CT-dendron conjugates decorated with G3-PEG(2000), G4-PEG(2000), and G5-PEG(1000), withthe latter two structures offering the best combination of sievingratio and small molecule activity.