Investigation of Nanolipoprotein Particles Entrapped Within Nanoporous Silica: A Novel Platform for Immobilization of Integral Membrane Proteins

Abstract

Immobilization of integral membrane proteins (IMPs) in transparent, nanoporous silica gels has proven to be a challenge, as current and previous techniques utilize liposomes as biological membrane hosts. The instability of liposomes in nanoporous gels is attributed by their size (∼150 nm) and altered structure and lipid dynamics upon entrapment within the nanometer scale pores (5-50 nm) of silica gel. This ultimately results in disruption of protein activity. We intend to overcome these barriers by using nanolipoprotein particles (NLPs) as biomembrane hosts. NLPs are discoidal patches of lipid bilayer that are belted by amphiphilic scaffold proteins and have an average thickness of 5 nm, with diameters ranging from 10-15 nm. The IMP-NLP complexes are synthesized in a cell-free environment, which circumvents traditional protein reconstitution in membranes. Bacteriorhodopsin - a robust IMP protein that indicates its proper conformation via distinct purple coloration - will serve as a model IMP for this system. The spectral and physical properties of bacteriorhodopsin-NLPs entrapped within the gel are examined, as well as the phase behavior of the lipids within the NLP, to ensure proper functionality of the system. This bio-inorganic hybrid nanomaterial possesses a variety of viable applications. The success of this work could lead to the development of novel platforms in several areas, including high-throughput drug screening, chromatography, and biosensors.