A new Kunitz-type snake toxin family associated with an original
mode of interaction with the vasopressin 2 receptor.
Laura Droctové1, Justyna Ciolek1,
Christiane Mendre2, Amélia Chorfa2,
Paola Huerta1, Christelle Carvalo1,
Charlotte Gouin1, Manon Lancien1,
Guillaume Blanchet1, Gregory Upert1,
Edwin De Pauw3, Peggy Barbe1,
Mathilde Keck1, Gilles Mourier1,
Bernard Mouillac2, Servent Denis1,
Ricardo C. Rodríguez de la Vega4, Loïc
Quinton3, Nicolas Gilles1*
1 Université Paris Saclay, CEA, INRAE, Département Médicaments et
Technologies pour la Santé (DMTS), SIMoS, 91191 Gif-sur-Yvette, France
2 Institut de Génomique Fonctionnelle, Université de Montpellier, CNRS,
INSERM, 34094 Montpellier, France
3 Laboratory of Mass Spectrometry, MolSys Research Unit, University of
Liège, Liège, Belgium
4 Ecologie, Systematique Evolution. CNRS, AgroParisTech, Université
Paris-Saclay, 91400 Orsay, France
*to whom correspondence should be addressed:nicolas.gilles@cea.fr
Nicolas Gilles ORCID 0000-0001-6853-9764
Denis Servent ORCID 0000-0002-0774-1691
Authorship contribution statement
L.D, C.M., G.B., M.K., R.C.R.V., L.Q. and N.G. participated in research
design.
L.D., J.C., C.M., A.C., P.H., C.C., C.G., M.L., G.B., G.U., P.B., M.K.,
G.M., R.C.R.V. and L.Q. conducted experiments.
L.D., G.B., G.M. and L.Q. contributed new reagents and analytic tools.
L.D., C.M., G.B., M.K., R.C.R.V. and N.G. performed the data analysis.
B.M., D.S., R.C.R.V., L.Q. and N.G. wrote or contributed to the writing
of the manuscript.
Funding statement
This work was supported by the French Atomic and Alternative Energies.
Conflicts of interest statement
No conflict of interest
Data Availability Statement: No data have been shared.
Background and purpose. Venomous animals express numerous
Kunitz-type peptides. The mambaquaretin-1 (MQ1) identified from theDendroaspis angusticeps venom is the most selective antagonist of
the arginine-vasopressin V2 receptor (V2R) and the unique Kunitz-type
peptide active on a GPCR. We aimed to exploit other mamba venoms to
enlarge the V2R-Kunitz peptide family and gain insight into the MQ1
molecular mode of action.
Experimental approach. We used a bio-guided screening assay to
identify novel MQs and placed them phylogenetically. MQs were produced
by solid phase peptide synthesis and characterized in vitro by
binding and functional tests and in vivo by diuresis measurement
in rats.
Key results. Eight additional MQs were identified with
nanomolar affinities for the V2R, all antagonists. MQs form a new
subgroup in the Kunitz family, close to the V2R non-active dendrotoxins
and to 2 V2R active cobra toxins. Sequence comparison between active and
non-active V2R Kunitz peptides highlighted 5 specific V2R positions.
Four of them are involved in V2R activity and belong to the 2 large MQ1
loops. We finally determined that 8 positions, part of these 2 loops,
interact with the V2R. The variant MQ1-K39A showed higher affinity for
the hV2R but not for the rat V2R.
Conclusions and implications. A third function and mode of
action is now associated with the Kunitz-peptides. The number of MQ1
residues involved in V2R binding is large and may explain its absolute
selectivity. MQ1-K39A represents the first step in the improvement of
the MQ1 design for medicinal perspective.
Abbreviations.
MQ Mambaquaretin
V2R arginine-vasopressin type 2 receptor
BPTI bovine pancreatic trypsin inhibitor
3FT 3 finger-fold toxins
Keywords: Snake venom, Kunitz peptide, V2R, antagonist, proteomics,
peptide sequencing, peptide synthesis.