TY - JOUR
T1 - Surface effects on functional amyloid formation
AU - Dear, Alexander J.
AU - Meisl, Georg
AU - Taylor, Christopher G.
AU - Palmiero, Umberto Capasso
AU - Stubbe, Susanne Nordby
AU - Liu, Qian
AU - Arosio, Paolo
AU - Linse, Sara
AU - Knowles, Tuomas P.J.
AU - Andreasen, Maria
PY - 2024/8/3
Y1 - 2024/8/3
N2 - Functional amyloids formed by the protein FapC in Pseudomonas bacteria are key structural components of Pseudomonas biofilms, which mediate chronic infections and also contribute to antimicrobial resistance. Here, we combine kinetic experiments with mechanistic modelling to probe the role of surfaces in FapC functional amyloid formation. We find that nucleation of new fibrils is predominantly heterogeneous in vitro, being catalysed by reaction vessel walls but not by the air/water interface. Removal of such interfaces by using microdroplets greatly slows heterogeneous nucleation and reveals a hitherto undetected fibril surface-catalysed “secondary nucleation” reaction step. We tune the degree of catalysis by varying the interface chemistry of the reaction vessel and by adding nanoparticles with tailored surface properties that catalyse fibril nucleation. In so doing, we discover that the rate of nucleation is controlled predominantly by the strength with which FapC binds to the catalytic sites on the interface, and by its surface area. Surprisingly, neither primary nucleation rate nor catalytic site binding strength appear closely correlated to the charge and hydrophilicity of the interface. This indicates the importance of considering experimental design in terms of surface chemistry of the reaction container while also highlighting the notion that fibril nucleation during protein aggregation is a heterogeneous process.
AB - Functional amyloids formed by the protein FapC in Pseudomonas bacteria are key structural components of Pseudomonas biofilms, which mediate chronic infections and also contribute to antimicrobial resistance. Here, we combine kinetic experiments with mechanistic modelling to probe the role of surfaces in FapC functional amyloid formation. We find that nucleation of new fibrils is predominantly heterogeneous in vitro, being catalysed by reaction vessel walls but not by the air/water interface. Removal of such interfaces by using microdroplets greatly slows heterogeneous nucleation and reveals a hitherto undetected fibril surface-catalysed “secondary nucleation” reaction step. We tune the degree of catalysis by varying the interface chemistry of the reaction vessel and by adding nanoparticles with tailored surface properties that catalyse fibril nucleation. In so doing, we discover that the rate of nucleation is controlled predominantly by the strength with which FapC binds to the catalytic sites on the interface, and by its surface area. Surprisingly, neither primary nucleation rate nor catalytic site binding strength appear closely correlated to the charge and hydrophilicity of the interface. This indicates the importance of considering experimental design in terms of surface chemistry of the reaction container while also highlighting the notion that fibril nucleation during protein aggregation is a heterogeneous process.
UR - http://www.scopus.com/inward/record.url?scp=85201273814&partnerID=8YFLogxK
U2 - 10.1039/d4nr01496k
DO - 10.1039/d4nr01496k
M3 - Journal article
C2 - 39135495
AN - SCOPUS:85201273814
SN - 2040-3364
VL - 16
SP - 16172
EP - 16182
JO - Nanoscale
JF - Nanoscale
IS - 34
ER -