High Throughput Methods for Discovering Membrane Active Peptides

Abstract

Rational design and engineering of membrane active peptides remains a largely unsatisfied goal. We have hypothesized that this us due, in part, to the fact that some membrane activities, such as permeabilization, and cell penetrating ability are not dependent on specific amino acid sequences or specific three-dimensional peptide structures. Instead they depend on interfacial activity; the ability of a molecule to partition into in the membrane-water interface and to strain the packing and organization of lipids. We are testing that idea by taking a novel approach to biomolecular engineering and design of membrane-active peptides. Several rational combinatorial peptide libraries containing 10-16,000 members have been screened for water soluble members that either permeabilize phospholipid membranes or translocate without permeabilization. Stringent, two-phase, high-throughput screens were used to identify dozens of unique peptides that had potent membrane permeabilizing activity or cell penetrating activity, but were also highly water soluble. These rare and uniquely active peptides did not always share a particular sequence motif, peptide length or net charge, but always share common compositional features, secondary structure and core hydrophobicity. We suggest that they function by common mechanisms that depends mostly on interfacial activity. We demonstrate here that composition-space peptide libraries coupled with function-based high-throughput screens can lead to the discovery of diverse, soluble, and highly potent interfacially active peptides.