Reconfigurable Peptide Analogs of Apolipoprotein A-I Reveal Tunable Features of Nanodisc Assembly

Nanodisc (ND)-forming membrane scaffold proteins or peptides developed from apolipoprotein A-I (apoA-I) have led to considerable promise in structural biology and therapeutic applications. However, the rationale and regularity characteristics in peptide sequence design remain inconclusive. Here, we proposed a consensus-based normalization approach through the reversed engineering of apoA-IΔ1-45 to design reconfigurable apoA-I peptide analogs (APAs) for tunable ND assembly. We present extensive morphological validations and computational simulation analyses on divergent APA-NDs that are generated by our method. Fifteen divergent APAs were generated accordingly to study the assembly machinery of NDs. We show that APA designs exhibit multifactorial influence in terms of varying APA tandem repeats, sequence composition, and lipid-to-APA ratio to form tunable diameters of NDs. There is a strong positive correlation between DMPC-to-APA ratios and ND diameters. Longer APA with more tandem repeats tends to yield higher particle size homogeneity. Our results also suggest proline is a dispensable residue for the APA-ND formation. Interestingly, proline-rich substitution not only provides an inward-bending effect in forming smaller NDs but also induces the cumulative chain flexibility that enables larger ND formation at higher lipid ratios. Additionally, proline-tryptophan residues in APAs play a dominant role in forming larger NDs. Molecular simulation shows that enriched basic and acidic residues in APAs evoke abundant hydrogen-bond and salt bridge networks to reinforce the structural stability of APA-NDs. Together, our findings provide a rational basis for understanding APA design. The proposed model could be extended to other apolipoproteins for desired ND engineering.