The effect of hydrophobic alkyl sidechains on size and solution behaviors of nanodiscs formed by alternating styrene maleamic copolymer

The development of amphipathic polymers, including various formulations of styrene-maleic acid (SMA) co-polymers, has allowed the purification of increasing sizes and complexities of biological membrane protein assemblies in native nanodiscs. However, the factors determining the sizes and shapes of the resulting bio-nano particles remain unclear. Here, we show how grafting on short alkyl amine sidechains onto the polar residues leads to a broad set of nanoparticle sizes with improved solution behavior. The solubilization of lipid vesicles occurs over a wide range of pH levels and calcium concentrations, providing utility across the physiologically relevant range of solution conditions. Furthermore, the active SMA derivatives can contain strictly alternating monomers, which have inherently lower sequence polydispersity. Pronounced differences in the shapes of native nanoparticles were formed from Escherichia coli bacterial outer membrane containing PagP protein using methyl, ethyl and propylamine derivatives of styrene-maleic anhydride. In particular, the methylamine-substituted polymer forms smaller, mono-dispersed nanodiscs, while the longer alkyl derivatives form worm-like nanostructures. Thus the introduction of hydrophobicity onto the polar sidechains of amphipathic polymers has profound effects on morphology of native nanodisc, with shorter methyl moieties offering more uniformity and utility for structural biology studies.