TY - JOUR
T1 - Functional aggregation of cell-free proteins enables fungal ice nucleation
AU - Schwidetzky, Ralph
AU - Ribeiro, Ingrid de Almeida
AU - Bothen, Nadine
AU - Backes, Anna T.
AU - DeVries, Arthur L.
AU - Bonn, Mischa
AU - Fröhlich-Nowoisky, Janine
AU - Molinero, Valeria
AU - Meister, Konrad
PY - 2023/11/14
Y1 - 2023/11/14
N2 - Biological ice nucleation plays a key role in the survival of cold-adapted organisms. Several species of bacteria, fungi, and insects produce ice nucleators (INs) that enable ice formation at temperatures above −10 °C. Bacteria and fungi produce particularly potent INs that can promote water crystallization above −5 °C. Bacterial INs consist of extended protein units that aggregate to achieve superior functionality. Despite decades of research, the nature and identity of fungal INs remain elusive. Here, we combine ice nucleation measurements, physicochemical characterization, numerical modeling, and nucleation theory to shed light on the size and nature of the INs from the fungus Fusarium acuminatum. We find ice-binding and ice-shaping activity of Fusarium IN, suggesting a potential connection between ice growth promotion and inhibition. We demonstrate that fungal INs are composed of small 5.3 kDa protein subunits that assemble into ice-nucleating complexes that can contain more than 100 subunits. Fusarium INs retain high ice-nucleation activity even when only the ~12 kDa fraction of size-excluded proteins are initially present, suggesting robust pathways for their functional aggregation in cell-free aqueous environments. We conclude that the use of small proteins to build large assemblies is a common strategy among organisms to create potent biological INs.
AB - Biological ice nucleation plays a key role in the survival of cold-adapted organisms. Several species of bacteria, fungi, and insects produce ice nucleators (INs) that enable ice formation at temperatures above −10 °C. Bacteria and fungi produce particularly potent INs that can promote water crystallization above −5 °C. Bacterial INs consist of extended protein units that aggregate to achieve superior functionality. Despite decades of research, the nature and identity of fungal INs remain elusive. Here, we combine ice nucleation measurements, physicochemical characterization, numerical modeling, and nucleation theory to shed light on the size and nature of the INs from the fungus Fusarium acuminatum. We find ice-binding and ice-shaping activity of Fusarium IN, suggesting a potential connection between ice growth promotion and inhibition. We demonstrate that fungal INs are composed of small 5.3 kDa protein subunits that assemble into ice-nucleating complexes that can contain more than 100 subunits. Fusarium INs retain high ice-nucleation activity even when only the ~12 kDa fraction of size-excluded proteins are initially present, suggesting robust pathways for their functional aggregation in cell-free aqueous environments. We conclude that the use of small proteins to build large assemblies is a common strategy among organisms to create potent biological INs.
KW - heterogeneous ice nucleation
KW - ice-nucleating proteins
KW - fungi
KW - protein assembly
U2 - 10.1073/pnas.2303243120
DO - 10.1073/pnas.2303243120
M3 - Article
C2 - 37943838
SN - 0027-8424
VL - 120
JO - Proceedings of the National Academy of Sciences
JF - Proceedings of the National Academy of Sciences
IS - 46
M1 - e2303243120
ER -