Maturation of the functional mouse CRES amyloid from globular form

Aveline Hewetson, Nazmul H. Khan, Matthew J. Dominguez, Hoa Quynh Do, R. E. Kusko, Collin G. Borcik, Daniel J. Rigden, Ronan M. Keegan, R. Bryan Sutton, Michael P. Latham, Benjamin J. Wylie, Gail A. Cornwall

Research output: Contribution to journalArticlepeer-review

2 Scopus citations

Abstract

The epididymal lumen contains a complex cystatin-rich nonpathological amyloid matrix with putative roles in sperm maturation and sperm protection. Given our growing understanding for the biological function of this and other functional amyloids, the problem still remains: how functional amyloids assemble including their initial transition to early oligomeric forms. To examine this, we developed a protocol for the purification of nondenatured mouse CRES, a component of the epididymal amyloid matrix, allowing us to examine its assembly to amyloid under conditions that may mimic those in vivo. Herein we use X-ray crystallography, solution-state NMR, and solid-state NMR to follow at the atomic level the assembly of the CRES amyloidogenic precursor as it progressed from monomeric folded protein to an advanced amyloid. We show the CRES monomer has a typical cystatin fold that assembles into highly branched amyloid matrices, comparable to those in vivo, by forming β-sheet assemblies that our data suggest occur via two distinct mechanisms: a unique conformational switch of a highly flexible disulfide-anchored loop to a rigid β-strand and by traditional cystatin domain swapping. Our results provide key insight into our understanding of functional amyloid assembly by revealing the earliest structural transitions from monomer to oligomer and by showing that some functional amyloid structures may be built by multiple and distinctive assembly mechanisms.

Original languageEnglish
Pages (from-to)16363-16372
Number of pages10
JournalProceedings of the National Academy of Sciences of the United States of America
Volume117
Issue number28
DOIs
StatePublished - Jul 14 2020

Keywords

  • Amyloid
  • Cystatin
  • NMR spectroscopy
  • X-ray crystallography

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