PT  - JOURNAL ARTICLE
AU  - David Pantoja-Uceda
AU  - Javier Oroz
AU  - Cristina Fernández
AU  - Eva de Alba
AU  - Rafael Giraldo
AU  - Douglas V. Laurents
TI  - Conformational Priming of RepA-WH1 for Functional Amyloid Conversion Detected by NMR Spectroscopy
AID  - 10.1101/612135
DP  - 2019 Jan 01
TA  - bioRxiv
PG  - 612135
4099  - http://biorxiv.org/content/early/2019/04/17/612135.short
4100  - http://biorxiv.org/content/early/2019/04/17/612135.full
AB  - How proteins with a stable globular fold acquire the amyloid state is still largely unknown. RepA is a versatile plasmidic DNA binding protein, functional either as a transcriptional repressor or as an initiator or inhibitor of DNA replication, the latter through the assembly of an amyloidogenic oligomer. Its N-terminal domain (WH1) is responsible for discrimination between these functional abilities by undergoing hitherto unknown structural changes. Furthermore, when expressed alone, RepA-WH1 behaves as a synthetic prion-like protein causing an amyloid proteinopathy in bacteria. RepA-WH1 is a stable dimer whose conformational dynamics had not been explored. Here we have studied it through NMR {1H}-15N relaxation and H/D exchange kinetics measurements. The N- and the C- terminal α-helices, which lock the WH1 fold in each subunit of the dimer, as well as an internal amyloidogenic loop, show reduced stability and are partially unfolded in solution. S4-indigo, a small molecule ligand known to interfere with the amyloid assembly of RepA-WH1, binds to and tethers the N-terminal α-helix and a β-hairpin that is involved in dimerization, thus providing evidence for a priming role of fraying ends and dimerization switches in the amyloidogenesis of folded proteins.