Design of Beta-2 Microglobulin Adsorbent Protein Nanoparticles

Abstract

Beta-2 microglobulin (B2M) is an immune system protein that is found on the surface of all nucleated human cells. B2M is naturally shed from cell surfaces into the plasma, followed by renal excretion. In patients with impaired renal function, B2M will accumulate in organs and tissues leading to significantly reduced life expectancy and quality of life. While current hemodialysis methods have been successful in managing electrolyte as well as small and large molecule disturbances arising in chronic renal failure, they have shown only modest success in managing plasma levels of B2M and similar sized proteins, while sparing important proteins such as albumin. We describe a systematic protein design effort aimed at adding the ability to selectively remove specific, undesired waste proteins such as B2M from the plasma of chronic renal failure patients. A novel nanoparticle built using a tetrahedral protein assembly as a scaffold that presents 12 copies of a B2M-binding nanobody is described. The designed nanoparticle binds specifically to B2M through protein–protein interactions with nanomolar binding affinity (~4.2 nM). Notably, binding to the nanoparticle increases the effective size of B2M by over 50-fold, offering a potential selective avenue for separation based on size. We present data to support the potential utility of such a nanoparticle for removing B2M from plasma by either size-based filtration or by polyvalent binding to a stationary matrix under blood flow conditions. Such applications could address current shortcomings in the management of problematic mid-sized proteins in chronic renal failure patients.

Designed fluorescent protein cages as fiducial markers for targeted cell imaging

“…Protein cages are hollow, nanoscale structures that can be designed to spontaneously self-assemble from multiple copies of modular protein subunits [11][12][13][14][15][16][17] . The resulting cage structures have well-defined sizes and shapes and can be engineered to encapsulate guest molecules, display functional proteins on their surfaces, and work as imaging scaffolds for cryo-electron microscopy (cryo-EM) [18][19][20][21][22][23][24][25][26] .…”

Design of a symmetry‐broken tetrahedral protein cage by a method of internal steric occlusion

“…For example, the outer surface of a symmetric protein cage will present many structurally and chemically equivalent motifs (e.g., many equivalent chain termini) for attachment or fusion—12 for symmetry T, 24 for symmetry O, and 60 for symmetry I. That feature is beneficial for some applications (e.g., vaccine‐like particles (Antanasijevic et al, 2020; Brouwer et al, 2021; Marcandalli et al, 2019; Walls et al, 2020), polyvalent binding (Divine et al, 2021; Miller et al, 2023), enzymatic materials (McConnell et al, 2020; McNeale et al, 2023), and imaging scaffolds (Castells‐Graells et al, 2023; Liu et al, 2018; Liu et al, 2019)). However, for other applications it may be desirable to functionalize (or present fusions) on a singular location.…”

A Suite of Designed Protein Cages Using Machine Learning Algorithms and Protein Fragment-Based Protocols

“…Achievements over the last decade have demonstrated the potential value of engineered protein cages and related types of SCMs for applications across the areas of nanotechnology and medicine. Their biocompatibility as well as their size, topology, and multivalency have enabled applications such as the localization of target substrates 10 , molecular delivery 11 or sequestration of payloads 12 , and scaffolding of antigens [13][14][15] , enzymes 16,17 , or binders for high resolution imaging 18,19 . Notwithstanding these promising demonstrations, successful designs have only scratched the surface with respect to the rich functional complexity and dynamics that are possible with protein assemblies, as exemplified by naturally evolved systems [20][21][22][23][24][25] .…”