Interaction of Synthetic Human SLURP-1 with the Nicotinic Acetylcholine Receptors

Abstract

Human SLURP-1 is a secreted protein of the Ly6/uPAR/three-finger neurotoxin family that co-localizes with nicotinic acetylcholine receptors (nAChRs) and modulates their functions. Conflicting biological activities of SLURP-1 at various nAChR subtypes have been based on heterologously produced SLURP-1 containing N- and/or C-terminal extensions. Here, we report the chemical synthesis of the 81 amino acid residue human SLURP-1 protein, characterization of its 3D structure by NMR, and its biological activity at nAChR subtypes. Radioligand assays indicated that synthetic SLURP-1 did not compete with [125I]-α-bungarotoxin (α-Bgt) binding to human neuronal α7 and Torpedo californica muscle-type nAChRs, nor to mollusk acetylcholine binding proteins (AChBP). Inhibition of human α7-mediated currents only occurred in the presence of the allosteric modulator PNU120596. In contrast, we observed robust SLURP-1 mediated inhibition of human α3β4, α4β4, α3β2 nAChRs, as well as human and rat α9α10 nAChRs. SLURP-1 inhibition of α9α10 nAChRs was accentuated at higher ACh concentrations, indicating an allosteric binding mechanism. Our results are discussed in the context of recent studies on heterologously produced SLURP-1 and indicate that N-terminal extensions of SLURP-1 may affect its activity and selectivity on its targets. In this respect, synthetic SLURP-1 appears to be a better probe for structure-function studies.

Makaluvamine G from the Marine Sponge Zyzzia fuliginosa Inhibits Muscle nAChR by Binding at the Orthosteric and Allosteric Sites

“…In summary, Makaluvamine G might present a potentially useful hit molecule for the rational design of drugs acting like un-competitive antagonists of nAChRs and other Cys-loop receptors. This might be of special interest in the light of findings that endogenous nAChR ligands from the Ly6 family show some properties similar to un-competitive inhibitors [ 30 , 31 , 32 ]. Thus, small molecules mimicking properties of endogenous modulators of Cys-loop receptors could be of a high value.…”

Oligoarginine Peptides, a New Family of Nicotinic Acetylcholine Receptor Inhibitors

“…In our recent work on low molecular weight compounds (Kudryavtsev et al, 2018;Spirova et al, 2019), a-conotoxins (Kudryavtsev et al, 2015;Kasheverov et al, 2016;Kryukova et al, 2018a), and human Ly6 proteins (Lyukmanova et al, 2016;Durek et al, 2017), we found that some of them can bind to the orthosteric or allosteric sites of nAChRs. Here we did not plan a detailed investigation of the mechanism of oligoarginine action, but we made exceptions for R8 and W2R4.…”

New Binding Mode of SLURP Protein to a7 Nicotinic Acetylcholine Receptor Revealed by Computer Simulations

“…Among the latter is SLURP-1 functioning as a water-soluble paracrine/autocrine messenger molecule which binds nicotinic acetylcholine receptors and regulates keratinocyte growth and angiogenesis. Such properties of SLURP-1 make it a valuable object to study as an endogenous cholinergic ligand similar to snake venom neurotoxins that played a crucial role in the nAChR research for decades [8,10].…”

“…Noteworthy, no experimental spatial structure of fusion SLURP-1 proteins were described. Fusion proteins demonstrated positive allosteric modulation of α7 nAChR while recombinant SLURP-1 act as an inhibitor of the receptor [8].…”