Structure and allosteric activity of a single-disulfide conopeptide from Conus zonatus at human α3β4 and α7 nicotinic acetylcholine receptors
Authors: Lotten Ragnarsson
Madhan Kumar Mohan
Benjamin Franklin Jayaseelan
Siddhartha P Sarma
Richard J Lewis
Rajesh R P
Nikita Abraham
Journal: Journal of Biological Chemistry (2020)
Discipline: BIOLOGY & BIOCHEMISTRY
Keywords: allosteric modulation; allosteric regulation; conformational exchange; disulfide; molecular pharmacology
Affiliations: Molecular Biophysics Unit, Indian Institute of Science, Bangalore, Karnataka 560012, India., Institute for Molecular Bioscience, Queensland Bioscience Precinct, The University of Queensland, 306 Carmody Rd., St. Lucia Queensland 4072, Australia., Sathyabama Institute of Science and Technology, Jeppiaar Nagar, Rajiv Gandhi Salai, Chennai 600119, Tamil Nadu, India., Bombay Natural History Society, Hornbill House, Dr. Salim Ali Chowk, Mumbai 400 001, Maharashtra, India., Institute for Molecular Bioscience, Queensland Bioscience Precinct, The University of Queensland, 306 Carmody Rd., St. Lucia Queensland 4072, Australia ., Molecular Biophysics Unit, Indian Institute of Science, Bangalore, Karnataka 560012, India .

Abstract:  Conopeptides are neurotoxic peptides in the venom of marine cone snails and have broad therapeutic potential for managing pain and other conditions. Here, we identified the single-disulfide peptides Czon1107 and Cca1669 from the venoms of Conus zonatus and Conus caracteristicus, respectively. We observed that Czon1107 strongly inhibits the human α3β4 (IC50 15.7 ± 3.0 μm) and α7 (IC50 77.1 ± 0.05 μm) nicotinic acetylcholine receptor (nAChR) subtypes, but the activity of Cca1669 remains to be identified. Czon1107 acted at a site distinct from the orthosteric receptor site. Solution NMR experiments revealed that Czon1107 exists in equilibrium between conformational states that are the result of a key Ser4-Pro5cis-trans isomerization. Moreover, we found that the X-Pro amide bonds in the inter-cysteine loop are rigidly constrained to cis conformations. Structure-activity experiments of Czon1107 and its variants at positions P5 and P7 revealed that the conformation around the X-Pro bonds (cis-trans) plays an important role in receptor subtype selectivity. The cis conformation at the Cys6-Pro7 peptide bond was essential for α3β4 nAChR subtype allosteric selectivity. In summary, we have identified a unique single-disulfide conopeptide with a noncompetitive, potentially allosteric inhibitory mechanism at the nAChRs. The small size and rigidity of the Czon1107 peptide could provide a scaffold for rational drug design strategies for allosteric nAChR modulation. This new paradigm in the "conotoxinomic" structure-function space provides an impetus to screen venom from other Conus species for similar, short bioactive peptides that allosterically modulate ligand-gated receptor function. 

Retracted: No

PubPeer comments:
 2020 correction. https: www.jbc.org content 295 26 8872.short There was an error in Fig. 5iv. The residue numbering in the spectrum should read as 8P and not 7P. The mislabeling was introduced during the revision stage. This error has now been corrected and does not affect the results or conclusions of this work. , 
 I do not see a comment. Only a reproduction of the correction issued by the authors and accepted by the journal. However, it appears that Digitalis ciliata (Why the pseudonym? artfully foxy though and hopefully not hand-in-glove) appears to want a response to an unarticulated question. Digitalis, I wish you had read the manuscript. I hope your emotions, upon coming across this manuscript, were not similar to those of let s say Groucho Marx who once said  From the moment I picked up your book until I put it down, I was convulsed with laughter. Some day I intend reading it or Dorothy Parker who said  I simply couldn t put this book down, i had to fling it with great force  . In any case, since I am at the receiving end, please find an explanation below. The study describes the activity and structure of a conopeptide and its variants , the sequences of which are given in Table 1 in the manuscript. It can be seen that in the peptide Czon1107-P7A, proline 7 has been replaced with alanine and that the peptide now has only two out of the three prolines present in the WT peptide. These prolines are at positions 5 and 8 and NOT AT 7!. Indeed the Figure legend does point to this. However, during the course of revising the manuscript, the spectrum was mislabeled as that belonging to P7 instead of P8. Figure revisions were sought by the reviewers journal included changes in font, removing extra tick marks on the axes etc. It was during this stage that the error was introduced. Earlier versions of the manuscript did have the correct labeling of the crosspeaks in the NMR spectrum and this can be verified by the journal. I would like to add that the NMR chemical shifts of aminoacids in the peptides have been deposited in the BMRB and which are given in TablesS-1D and E (supplementary information). In addition, the NMR spectra on page S7 (supplementary information) also clearly point to the sequence and chemical shift assignment of Czon1107-P7A. Lastly, the structures shown in Figure7i,ii also indicate that the peptides are indeed of Czon1107-P7A. I would like to end with the lines of a poem ``` So ripeness climbs the bells of Digitalis, flower by flower, undistracted by a Mind, or a Design, or by desire. - The Miracle of the Bees and the Foxgloves ANNE STEVENSON ``` Just a comment. When choosing to follow the Linnaeus system of nomenclature, the species is always written in lower case. ,