Miniaturized bioaffinity assessment coupled to mass spectrometry for guided purification of bioactives from toad and cone snail.
A nano-flow high-resolution screening platform, featuring a parallel chip-based microfluidic bioassay and mass spectrometry coupled to nano-liquid chromatography, was applied to screen animal venoms for nicotinic acetylcholine receptor like (nAChR) affinity by using the acetylcholine binding protein, a mimic of the nAChR. The potential of this microfluidic platform is demonstrated by profiling the Conus textile venom proteome, consisting of over 1,000 peptides. Within one analysis (<90 min, 500 ng venom injected), ligands are detected and identified. To show applicability for non-peptides, small molecular ligands such as steroidal ligands were identified in skin secretions from two toad species (Bufo alvarius and Bufo marinus). Bioactives from the toad samples were subsequently isolated by MS-guided fractionation. The fractions analyzed by NMR and a radioligand binding assay with α7-nAChR confirmed the identity and bioactivity of several new ligands.
AIMMS Division of BioMolecular Analysis, Faculty of Sciences, VU University Amsterdam, De Boelelaan 1081, 1083 HV Amsterdam, The Netherlands. ferryheus@gmail.com.
Ferry
F
Heus
AIMMS Division of BioMolecular Analysis, Faculty of Sciences, VU University Amsterdam, De Boelelaan 1081, 1083 HV Amsterdam, The Netherlands. r.a.otvos@vu.nl.
Reka A
RA
Otvos
Department of Biophysical Chemistry, Institute for Molecules and Materials, Radboud University Nijmegen, Heyendaalseweg 135, 6525 AJ Nijmegen, The Netherlands. r.aspers@nmr.ru.nl.
Ruud L E G
RL
Aspers
Department of Molecular and Cellular Neurobiology, Center for Neurogenomics and Cognitive Research, Neuroscience Campus Amsterdam, VU University Amsterdam, De Boelelaan 1085, 1081 HV Amsterdam, The Netherlands. rene.van.elk@vu.nl.
Rene
R
van Elk
AIMMS Division of BioMolecular Analysis, Faculty of Sciences, VU University Amsterdam, De Boelelaan 1081, 1083 HV Amsterdam, The Netherlands. j.i.halff@student.vu.nl.
Jenny I
JI
Halff
AIMMS Division of Organic Chemistry, Faculty of Sciences, VU University Amsterdam, De Boelelaan 1081, 1083 HV Amsterdam, The Netherlands. a.w.ehlers@vu.nl.
Andreas W
AW
Ehlers
The Institute for Molecular Bioscience, The University of Queensland, St Lucia, Queensland 4072, Australia. s.dutertre@imb.uq.edu.au.
Sébastien
S
Dutertre
The Institute for Molecular Bioscience, The University of Queensland, St Lucia, Queensland 4072, Australia. r.lewis@imb.uq.edu.au.
Richard J
RJ
Lewis
Department of Biophysical Chemistry, Institute for Molecules and Materials, Radboud University Nijmegen, Heyendaalseweg 135, 6525 AJ Nijmegen, The Netherlands. s.wijmenga@nmr.ru.nl.
Sybren
S
Wijmenga
Department of Molecular and Cellular Neurobiology, Center for Neurogenomics and Cognitive Research, Neuroscience Campus Amsterdam, VU University Amsterdam, De Boelelaan 1085, 1081 HV Amsterdam, The Netherlands. guus.smit@vu.nl.
August B
AB
Smit
AIMMS Division of BioMolecular Analysis, Faculty of Sciences, VU University Amsterdam, De Boelelaan 1081, 1083 HV Amsterdam, The Netherlands. w.m.a.niessen@vu.nl.
Wilfried M A
WM
Niessen
AIMMS Division of BioMolecular Analysis, Faculty of Sciences, VU University Amsterdam, De Boelelaan 1081, 1083 HV Amsterdam, The Netherlands. j.kool@vu.nl.
Jeroen
J
Kool