Role of backbone strain in de novo design of complex \u03b1/\u03b2 protein structures

Nobuyasu Koga,David Baker,Javier Castellanos,Gaetano T Montelione,Gaohua Liu,Rie Koga

doi:10.1038/s41467-021-24050-7

Abstract

We previously elucidated principles for designing ideal proteins with completely consistent local and non-local interactions which have enabled the design of a wide range of new αβ-proteins with four or fewer β-strands. The principles relate local backbone structures to supersecondary-structure packing arrangements of α-helices and β-strands. Here, we test the generality of the principles by employing them to design larger proteins with five- and six- stranded β-sheets flanked by α-helices. The initial designs were monomeric in solution with high thermal stability, and the nuclear magnetic resonance (NMR) structure of one was close to the design model, but for two others the order of strands in the β-sheet was swapped. Investigation into the origins of this strand swapping suggested that the global structures of the design models were more strained than the NMR structures. We incorporated explicit consideration of global backbone strain into the design methodology, and succeeded in designing proteins with the intended unswapped strand arrangements. These results illustrate the value of experimental structure determination in guiding improvement of de novo design, and the importance of consistency between local, supersecondary, and global tertiary interactions in determining protein topology. The augmented set of principles should inform the design of larger functional proteins.

Highlights

We previously elucidated principles for designing ideal proteins with completely consistent local and non-local interactions which have enabled the design of a wide range of new αβ-proteins with four or fewer β-strands
We selected as design targets two topologies which are widespread in enzymes in nature: the P-loop fold and the Rossmann fold
We considered the second possibility more likely because the two strands which swap are internal to the β-sheet, and have very similar patterns of hydrophobic residues; the sidechain–sidechain interactions in the design model and the nuclear magnetic resonance (NMR) structure are similar

Summary

Introduction

We previously elucidated principles for designing ideal proteins with completely consistent local and non-local interactions which have enabled the design of a wide range of new αβ-proteins with four or fewer β-strands. We incorporated explicit consideration of global backbone strain into the design methodology, and succeeded in designing proteins with the intended unswapped strand arrangements These results illustrate the value of experimental structure determination in guiding improvement of de novo design, and the importance of consistency between local, supersecondary, and global tertiary interactions in determining protein topology. There has been considerable progress in de novo protein design, stemming in part from the elucidation of principles[6,18] for designing ideal protein structures[19] stabilized by consistent local and nonlocal interactions These principles are embodied in a set of design rules relating local backbone structures to supersecondary structure packing of α-helices on paired β-strands, which generate funnel-shaped energy landscapes by disfavoring non-native states[6,9]. Investigation into the origins of this strand swapping revealed that the design principles must be extended to incorporate explicit consideration of global backbone strain to provide control over folded topologies for larger αβ-proteins

Methods

Results

Conclusion