title: Direct quantitation of peptide-mediated protein transport across a droplet-interface bilayer.
abstract: We introduce a new method for monitoring and quantitating the transport of materials across a model cell membrane. As a proof-of-concept, the cell-penetrating peptide, Pep-1, was used to carry horseradish peroxidase (HRP) across droplet-interface bilayers (DIBs). Two submicroliter, lipid-encased aqueous droplets form a membrane at the contacting interface, through which enzyme-peptide complexes pass during transport. Following transport, the droplets are separated and the captured enzymes are assayed by a fluorogenic reaction. The DIB method recapitulates the findings of earlier studies involving Pep-1, including the dependence of protein transport on voltage and membrane charge, while also contributing new insights. Specifically, we found that leaflet charge symmetry may play a role in Pep-1-mediated protein translocation. We anticipate that the DIB method may be useful for a variety of transport-based studies.
authors.affiliation: Department of Chemistry, University of Massachusetts, 710 North Pleasant Street, Amherst, Massachusetts 01003, United States.
authors.forename: Jing
authors.initials: J
authors.lastname: Huang
authors.forename: Max
authors.initials: M
authors.lastname: Lein
authors.forename: Christopher
authors.initials: C
authors.lastname: Gunderson
authors.forename: Matthew A
authors.initials: MA
authors.lastname: Holden
mesh_terms: D001671:Biochemistry / Q000379:methods; D003543:Cysteamine / Q000031:analogs & derivatives* / Q000378:metabolism; D006735:Horseradish Peroxidase / Q000008:administration & dosage; D008051:Lipid Bilayers / Q000378:metabolism*; D010455:Peptides / Q000378:metabolism*; D021381:Protein Transport
chemical_list: D008051:Lipid Bilayers; C503924:Pep-1 peptide; D010455:Peptides; D003543:Cysteamine; D006735:Horseradish Peroxidase