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Biophysics of Proteins at Surfaces: Assembly, Activation, Signaling
Tuesday Speaker Abstracts
Self-Assembly of Protein Nanofibrils that Display Active Enzymes
Sarah Perrett
.
Institute of Biophysics, Chinese Academy of Sciences, Beijing, China.
The ability of proteins to self-assemble into beta-sheet-rich aggregates called amyloid fibrils is
considered to be universal, although certain polypeptide sequences have a particularly high
propensity to adopt these conformations. In many cases the formation of amyloid fibrils is
deleterious and associated with the progression of disease, but there are also examples of
proteins for which the cross-beta structure represents the functional conformation. Ure2 is the
protein determinant of the yeast prion [URE3]. Ure2 consists of an N-terminal prion-inducing
domain that is disordered in the native state, whereas the C-terminal functional domain has a
globular fold with structural similarity to glutathione transferase enzymes. The C-terminal
domain shows enzymatic activity in both the soluble and fibrillar forms of Ure2. We have used a
variety of biophysical approaches to investigate the structure of Ure2 fibrils and their mechanism
of assembly. We have also created chimeric constructs where the prion domain is genetically
fused to other enzymes of different sizes and architectures. These chimeric polypeptide
constructs spontaneously self-assemble into nanofibrils with fused active enzyme subunits
displayed on the amyloid fibril surface. We can measure steady-state kinetic parameters for the
appended enzymes in situ within fibrils, and compare these for the identical protein constructs in
solution. We have also applied microfluidic techniques to form enzymatically-active microgel
particles from the chimeric self-assembling protein nanofibrils. The use of scaffolds formed from
biomaterials that self-assemble under mild conditions enables the formation of catalytic
microgels whilst maintaining the integrity of the encapsulated enzyme. In combination with
microfluidic trapping techniques, these approaches illustrate the potential of self-assembling
materials for enzyme immobilization and recycling, and for biological flow-chemistry. The
design principles can be adopted to create countless other bioactive amyloid-based materials with
diverse functions.