Forisomes are giant self-assembling mechanoproteins that undergo reversible structural changes in response to Ca2+ and various other stimuli. Artificial forisomes assembled from the monomer MtSEO-F1 can be used as smart biomaterials, but the molecular basis of their functionality is not understood. To determine the role of protein polymerization in forisome activity, we tested the Ca2+ association of MtSEO-F1 dimers (the basic polymerization unit) by circular dichroism spectroscopy and microscale thermophoresis. We found that soluble MtSEO-F1 dimers neither associate with Ca2+ nor undergo structural changes. However, polarization modulation infrared reflection absorption spectroscopy revealed that aggregated MtSEO-F1 dimers and fully-assembled forisomes associate with Ca2+ , allowing the hydration of poorly-hydrated protein areas. A change in the signal profile of complete forisomes indicated that Ca2+ interacts with negatively-charged regions in the protein complexes that only become available during aggregation. We conclude that aggregation is required to establish the Ca2+ response of forisome polymers.
Keywords: SUMO tag; calcium association; circular dichroism; forisome; microscale thermophoresis; multi-angle light scattering; polarization modulation infrared reflection absorption spectroscopy; protein aggregation; protein dimerization; responsive polymer.
© 2021 The Authors. Protein Science published by Wiley Periodicals LLC on behalf of The Protein Society.