RT Journal Article
SR Electronic
T1 Screening of hydrocarbon-stapled peptides for inhibition of calcium-triggered exocytosis
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2022.03.21.484632
DO 10.1101/2022.03.21.484632
A1 Ying Lai
A1 Michael J. Tuvim
A1 Jeremy Leitz
A1 John Peters
A1 Richard A. Pfuetzner
A1 Luis Esquivies
A1 Qiangjun Zhou
A1 Barbara Czako
A1 Jason B. Cross
A1 Philip Jones
A1 Burton F. Dickey
A1 Axel T. Brunger
YR 2022
UL http://biorxiv.org/content/early/2022/05/11/2022.03.21.484632.abstract
AB The so-called primary interface between the SNARE complex and synaptotagmin-1 (Syt1) is essential for Ca2+-triggered neurotransmitter release in neuronal synapses. The interacting residues of the primary interface are conserved across different species for synaptotagmins (Syt1, Syt2, Syt9), SNAP-25, and syntaxin-1A homologs involved in fast synchronous release. This Ca2+-independent interface forms prior to Ca2+-triggering and plays a role in synaptic vesicle priming. This primary interface is also conserved in the fusion machinery that is responsible for mucin granule membrane fusion. Ca2+-stimulated mucin secretion is mediated by the SNAREs syntaxin-3, SNAP-23, VAMP8, synaptotagmin-2, and other proteins. Here, we designed and screened a series of hydrocarbon-stapled peptides consisting of SNAP-25 fragments that included some of the key residues involved in the primary interface as observed in high-resolution crystal structures. We selected a subset of four stapled peptides that were highly α-helical as assessed by circular dichroism and that inhibited both Ca2+-independent and Ca2+-triggered ensemble lipid-mixing with neuronal SNAREs and Syt1. In a single-vesicle contentmixing assay with reconstituted neuronal SNAREs and synaptotagmin-1 or with reconstituted airway SNAREs and synaptotagmin-2, the selected peptides also suppressed Ca2+-triggered fusion. Taken together, hydrocarbon-stapled peptides that interfere with the primary interface consequently inhibit Ca2+-triggered exocytosis. Our inhibitor screen suggests that these compounds may be useful to combat mucus hypersecretion that is a major cause of airway obstruction in the pathophysiology of COPD, asthma and cystic fibrosis.Competing Interest StatementThe authors have declared no competing interest.