A Mutant Complement Factor H (W1183R) Enhances Proteolytic Cleavage of von Willebrand Factor by ADAMTS13 Under Shear.

Abstract

Background: A loss-of-functional mutation (W1183R) in human complement factor H (CFH) is associated with complement-associated hemolytic uremic syndrome; mice carrying a similar mutation (W1206R) in CFH also develop thrombotic microangiopathy but its plasma von Willebrand factor (VWF) multimer sizes were dramatically reduced. The mechanism underlying such a dramatic change in plasma VWF multimer distribution in these mice is not fully understood.

Objective and methods: To determine the VWF and CFH interaction and how CFH proteins affect VWF multimer distribution, we employed recombinant protein expression, purification, and various biochemical and biophysical tools.

Results: Purified recombinant W1183R-CFH but not wild-type (WT) CFH protein enhanced the proteolytic cleavage of both peptidyl and multimeric VWF substrates by recombinant ADAMTS13 in a concentration-dependent manner. Microscale thermophoresis assay demonstrated that both W1183R-CFH and WT-CFH proteins bound various VWF fragments (e.g., AIM-A1, A1-A2-A3, D'D3, D'D3-A1, and D'D3-A1-A2) with high affinities. Optical tweezer experiments further showed a concentration-dependent alteration in the contour length (L_c) and the persistent length (L_p) following pulling VWF-A2 domain in the presence of W1183R-CFH or WT-CFH protein. AlphaFold experiments revealed conformational changes in the VWF-A2, particularly the central region where the cleavage bond resides following addition of W1183R-CFH or WT-CFH protein.

Conclusions: These results demonstrate for the first time that W1183R-CFH but not WT-CFH protein enhances the proteolytic cleavage of VWF by ADAMTS13 under shear. This may be achieved by mechanic-induced conformational changes of the central A2 domain, leading to an altered cleavage of Tyr¹⁶⁰⁵-Met¹⁶⁰⁶ bond by ADAMTS13 under pathophysiological conditions.