Mar. Drugs 2012, 10(7), 1605-1618; doi:10.3390/md10071605
Article

Functional Expression in Escherichia coli of the Disulfide-Rich Sea Anemone Peptide APETx2, a Potent Blocker of Acid-Sensing Ion Channel 3

Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD 4072, Australia These authors contributed equally to this work.
* Authors to whom correspondence should be addressed.
Received: 4 June 2012; in revised form: 18 July 2012 / Accepted: 19 July 2012 / Published: 23 July 2012
(This article belongs to the Special Issue Sea Anemone Toxins)
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Abstract: Acid-sensing ion channels (ASICs) are proton-gated sodium channels present in the central and peripheral nervous system of chordates. ASIC3 is highly expressed in sensory neurons and plays an important role in inflammatory and ischemic pain. Thus, specific inhibitors of ASIC3 have the potential to be developed as novel analgesics. APETx2, isolated from the sea anemone Anthopleura elegantissima, is the most potent and selective inhibitor of ASIC3-containing channels. However, the mechanism of action of APETx2 and the molecular basis for its interaction with ASIC3 is not known. In order to assist in characterizing the ASIC3-APETx2 interaction, we developed an efficient and cost-effective Escherichia coli periplasmic expression system for the production of APETx2. NMR studies on uniformly 13C/15N-labelled APETx2 produced in E. coli showed that the recombinant peptide adopts the native conformation. Recombinant APETx2 is equipotent with synthetic APETx2 at inhibiting ASIC3 channels expressed in Xenopus oocytes. Using this system we mutated Phe15 to Ala, which caused a profound loss of APETx2’s activity on ASIC3. These findings suggest that this expression system can be used to produce mutant versions of APETx2 in order to facilitate structure-activity relationship studies.
Keywords: ASIC3; APETx2; heterologous expression; NMR; E. coli

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MDPI and ACS Style

Anangi, R.; Rash, L.D.; Mobli, M.; King, G.F. Functional Expression in Escherichia coli of the Disulfide-Rich Sea Anemone Peptide APETx2, a Potent Blocker of Acid-Sensing Ion Channel 3. Mar. Drugs 2012, 10, 1605-1618.

AMA Style

Anangi R, Rash LD, Mobli M, King GF. Functional Expression in Escherichia coli of the Disulfide-Rich Sea Anemone Peptide APETx2, a Potent Blocker of Acid-Sensing Ion Channel 3. Marine Drugs. 2012; 10(7):1605-1618.

Chicago/Turabian Style

Anangi, Raveendra; Rash, Lachlan D.; Mobli, Mehdi; King, Glenn F. 2012. "Functional Expression in Escherichia coli of the Disulfide-Rich Sea Anemone Peptide APETx2, a Potent Blocker of Acid-Sensing Ion Channel 3." Mar. Drugs 10, no. 7: 1605-1618.

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