Poster Presentation The 46th Lorne Conference on Protein Structure and Function 2021

Re-designing pore structure in self-assembling protein cages (#217)

Nuren Tasneem 1 , Lachlan Adamson 1 , Michael P. Andreas 2 , William Close 3 , Taylor N. Szyszka 1 , Eric Jenner 1 , Reginald Young 1 , Li-Chen Cheah 4 , Alexander Norman 1 , Frank Sainsbury 4 , Tobias W. Giessen 2 5 , Yu Heng Lau 1 6
  1. School of Chemistry, University of Sydney, Sydney, NSW, Australia
  2. Department of Biomedical Engineering, University of Michigan Medical School, Ann Arbor, Michigan, USA
  3. Sydney Microscopy and Microanalysis, University of Sydney, Sydney, NSW, Australia
  4. School of Environment and Science, Griffith University, Southport, QLD, Australia
  5. Department of Biological Chemistry, University of Michigan Medical School, Ann Arbor, Michigan, USA
  6. Sydney Nano Institute, University of Sydney, Sydney, NSW, Australia

Composed entirely of protein subunits, protein cages are self-assembling structures that are found in a broad range of microorganisms. In nature, protein cages in the form of viral capsids and bacterial microcompartments perform a diverse array of functions – from encapsulating genomic material to coordinating metabolic pathways within the cell.1,2 Protein cages can also be engineered to serve as templates for synthetic organelles, reaction chambers and sensing devices.3–5 Central to such applications is the notion that these perforated cages can act as a selectively permeable barrier by regulating molecular flux at the pores.6 However, the factors controlling molecular recognition at the cage pores remain unexplored.

To construct stable and selectively permeable cage structures, we re-designed the pore architecture of a class of protein cages known as encapsulins. We used the encapsulin protein cage from the bacterium, Thermotoga maritima 7 and designed a 24-member variant library by altering the pore size and/or charge. We then systematically examined the effects of both the structure and chemical nature of the pore on overall cage stability and molecular flux of cations into the cage. We report on twelve designs that were successfully purified, of which eight were found to exhibit prolonged thermal stability. Together with seven cryo-EM structures of cage variants, here we shed light on how pore modifications can affect solubility and stability of protein cages - towards predictable re-design of encapsulin protein cage pores

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