02897nas a2200265   4500008004100000022001400041245010000055210006900155260001500224300001400239490000700253520194900260653002502209653003002234653004502264653001302309653002202322100002602344700002502370700002002395700002302415700002002438700002002458856015302478       2021                        eng d  a1526-460200aNon-Covalent Carrier Hydrophobicity as a Universal Predictor of Intracellular Protein Activity.0 aNonCovalent Carrier Hydrophobicity as a Universal Predictor of I  c2021 07 12  a2850-28630 v223 a<p>Over the past decade, extensive optimization of polymeric cell-penetrating peptide (CPP) mimics (CPPMs) by our group has generated a substantial library of broadly effective carriers which circumvent the need for covalent conjugation often required by CPPs. In this study, design rules learned from CPPM development were applied to reverse-engineer the first library of simple amphiphilic block copolypeptides for non-covalent protein delivery, namely, poly(alanine--arginine), poly(phenylalanine--arginine), and poly(tryptophan--arginine). This new CPP library was screened for enhanced green fluorescent protein and Cre recombinase delivery alongside a library of CPPMs featuring equivalent side-chain configurations. Due to the added hydrophobicity imparted by the polymer backbone as compared to the polypeptide backbone, side-chain functionality was not a universal predictor of carrier performance. Rather, overall carrier hydrophobicity predicted the top performers for both internalization and activity of protein cargoes, regardless of backbone identity. Furthermore, comparison of protein uptake and function revealed carriers which facilitated high gene recombination despite remarkably low Cre internalization, leading us to formalize the concept of intracellular availability (IA) of the delivered cargo. IA, a measure of cargo activity per quantity of cargo internalized, provides valuable insight into the physical relationship between cellular internalization and bioavailability, which can be affected by bottlenecks such as endosomal escape and cargo release. Importantly, carriers with maximal IA existed within a narrow hydrophobicity window, more hydrophilic than those exhibiting maximal cargo uptake. Hydrophobicity may be used as a scaffold-independent predictor of protein uptake, function, and IA, enabling identification of new, effective carriers which would be overlooked by uptake-based screening methods.</p>
10aBiological Transport10aCell-Penetrating Peptides10aHydrophobic and Hydrophilic Interactions10aPolymers10aProtein Transport1 aHango, Christopher, R1 aBacklund, Coralie, M1 aDavis, Hazel, C1 aPosey, Nicholas, D1 aMinter, Lisa, M1 aTew, Gregory, N  uhttps://www.umass.edu/veterinary-animal-sciences/research-faculty/publications/non-covalent-carrier-hydrophobicity-universal-predictor-intracellular