Isolation and Pharmacological Characterization of α-Elapitoxin-Ot1a, a Short-Chain Postsynaptic Neurotoxin from the Venom of the Western Desert Taipan, Oxyuranus temporalis
Abstract
:1. Introduction
2. Results
2.1. Fractionation of Venom via Reverse-Phase HPLC
2.2. Intact Protein Analysis with MALDI-TOF Mass Spectrometry
2.3. Identification and de Novo Sequencing by LCMS/MS
2.4. In Vitro Neurotoxicity
2.5. CCh Cumulative Concentration-Response Curves
3. Discussion
4. Materials and Methods
4.1. Venom Supply
4.2. Purification α-Elapitoxin-Ot1a
4.3. Mass Spectrometry and Amino Acid Sequencing
4.3.1. Intact Protein Analysis Using Matrix Associated Laser Desorption Time of Flight (MALDI-TOF) Mass Spectrometry
4.3.2. In-Solution Digestion of Sample in Preparation for Electrospray-Ionization Coupled with Mass-Spectrometry/Mass Spectrometry (ESI-LCMS/MS)
4.3.3. Nanoflow Liquid Chromatography ESI-LCMS/MS
4.3.4. Protein Identification by Automated de Novo Sequencing
4.4. Neurotoxicity Studies
4.5. CCh Cumulative Concentration-Response Curves
4.6. Data Analysis
4.7. Chemicals and Drugs
Acknowledgments
Author Contributions
Conflicts of Interest
Abbreviations
ACh | acetylcholine |
CCh | carbachol |
HPLC | high performance liquid chromatography |
ESI-LCMS/MS | electrospray-ionization coupled with mass-spectrometry/mass spectrometry |
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Species | α-Neurotoxin | MW | Partial/Full N-Terminal Sequence |
---|---|---|---|
O. temporalis | α-elapitoxin-Ot1a a | 6712 | MTCYNQQSSQ AKTTTTCSGG VSSCYRKTWS DTRGTIIERG CGCPSVKKGI ERICCGTDKC NN |
O. scutellatus | α-scutoxin 1 b | 6781 | MTCYNQQSSE AKTTTTCSGG VSSCYKKTWY DGRGTRIERG |
O. scutellatus | α-oxytoxin 1 b | 6770 | MTCYNQQSSE AKTTTTCSGG VSSCYKETWY DGRGTT |
O. microlepidotus | oxylepitoxin-1 c | 6789 | MTCYNQQSSE AKTTTTCSGG VSSCYKETWY |
O. scutellatus | Taipan toxin 1 d | 6726 | MTCYNQQSSE AKTTTTCSGG VSSCYKKTWS DGRGTIIERG CGCPSVKKGI ERICCRTDKC NN |
O. scutellatus | Taipan toxin 2 d | 6781 | MTCYNQQSSE AKTTTTCSGG VSSCYKKTWS DIRGTIIERG CGCPSVKKGI ERICCRTDKC NN |
P. colletti | α-elapitoxin-Pc1a e | 6759.6 | MTCCNQQSSQ PKTTTTCAGG ETSCYKKTWS DHRGSRTERG CGCPHVKPGI KLTCCKTDEC NN |
P. porphyriacus | α-elapitoxin-Ppr1 e | 6746.5 | MTCCNQQSSQ PKTTTTCAGG ESSCYKKTWS DHRGSRTERG CGCPHVKPGI KLTCCETDEC NN |
P. papuanus | Papuantoxin-1 f | 6738 | MTCCNQQSSQ PKTTTT |
H. stephensi | Hostoxin-1 g | 6660 | MTPCNQQSSQ PKTTK |
Species | Common Name | Postsynaptic Neurotoxin | t90 (min) at 1 µM |
---|---|---|---|
O. temporalis | Western Desert Taipan | α-elapitoxin-Ot1a | 9.83 ± 0.36 a |
O. scutellatus | Coastal Taipan | α-scutoxin 1 | ~12 b |
O. scutellatus | Papuan Taipan | α-oxytoxin 1 | ~25 b |
O. microlepidotus | Inland Taipan | oxylepitoxin-1 | ~55 c |
H. stephensi | Stephen’s Banded snake | hostoxin-1 | ~10 d |
A. sp. Seram | Seram death adder | Acantoxin IVa | 9.7 ± 1.1 e |
Snake Species | Common Name | Toxin | pA2 Value |
---|---|---|---|
Bungarus multicinctus | Various kraits | α-bungarotoxin | 8.71 ± 0.06 a |
Hoplocephalus stephensi | Stephen’s Banded snake | hostoxin-1 | 8.45 ± 0.32 b |
Oxyuranus scutellatus | Coastal Taipan | α-scutoxin 1 | 8.38 ± 0.59 c |
Acanthophis sp. Seram | Seram death adder | acantoxin Iva | 8.36 ± 0.17 a |
Oxyuranus temporalis | Western Desert Taipan | α-elapitoxin-Ot1a | 8.02 ± 0.05 * |
Oxyuranus scutellatus | Papuan Taipan | α-oxytoxin 1 | 7.62 ± 0.04 c |
Oxyuranus microlepidotus | Inland Taipan | oxylepitoxin-1 | 7.16 ± 0.28 d |
Pseudechis colletti | Collett’s snake | α -elapitoxin-Pc1 | 7.04 ± 0.07 e |
Pseudechis prophyriacus | Red bellied black snake | α-elapitoxin-Ppr1 | 6.97 ± 0.03 e |
Pseudechis papuanus | Papuan black snake | papuantoxin-1 | 6.90 ± 0.30 f |
Not applicable | - | d-tubocurarine | 6.29 ± 0.06 a |
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Barber, C.M.; Ahmad Rusmili, M.R.; Hodgson, W.C. Isolation and Pharmacological Characterization of α-Elapitoxin-Ot1a, a Short-Chain Postsynaptic Neurotoxin from the Venom of the Western Desert Taipan, Oxyuranus temporalis. Toxins 2016, 8, 58. https://doi.org/10.3390/toxins8030058
Barber CM, Ahmad Rusmili MR, Hodgson WC. Isolation and Pharmacological Characterization of α-Elapitoxin-Ot1a, a Short-Chain Postsynaptic Neurotoxin from the Venom of the Western Desert Taipan, Oxyuranus temporalis. Toxins. 2016; 8(3):58. https://doi.org/10.3390/toxins8030058
Chicago/Turabian StyleBarber, Carmel M., Muhamad Rusdi Ahmad Rusmili, and Wayne C. Hodgson. 2016. "Isolation and Pharmacological Characterization of α-Elapitoxin-Ot1a, a Short-Chain Postsynaptic Neurotoxin from the Venom of the Western Desert Taipan, Oxyuranus temporalis" Toxins 8, no. 3: 58. https://doi.org/10.3390/toxins8030058
APA StyleBarber, C. M., Ahmad Rusmili, M. R., & Hodgson, W. C. (2016). Isolation and Pharmacological Characterization of α-Elapitoxin-Ot1a, a Short-Chain Postsynaptic Neurotoxin from the Venom of the Western Desert Taipan, Oxyuranus temporalis. Toxins, 8(3), 58. https://doi.org/10.3390/toxins8030058