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Mar. Drugs 2016, 14(1), 11; doi:10.3390/md14010011

Recombinant Expression and Characterization of α-Conotoxin LvIA in Escherichia coli

1
Key Laboratory of Tropical Biological Resources, Ministry of Education, Key Laboratory for Marine Drugs of Haikou, Hainan University, Haikou 570228, China
2
College of Horticulture and Landscapes, Hainan University, Haikou 570228, China
3
College of Marine Science, Hainan University, Haikou 570228, China
*
Author to whom correspondence should be addressed.
Academic Editor: Paul Long
Received: 13 October 2015 / Revised: 11 December 2015 / Accepted: 28 December 2015 / Published: 5 January 2016
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Abstract

α-Conotoxin LvIA is derived from Conus lividus, native to Hainan, and is the most selective inhibitor of α3β2 nicotinic acetylcholine receptors (nAChRs) known to date. In this study, an efficient approach for the production of recombinant α-Conotoxin LvIA is described. Tandem repeats of a LvIA gene fragment were constructed and fused with a KSI gene and a His6 tag in a Escherichia coli (E. coli) expression vector pET-31b(+). The recombinant plasmids were transformed into E. coli and were found to express well. The KSI-(LvIA)n-His6 fusion protein was purified by metal affinity chromatography and then cleaved with CNBr to release recombinant LvIA (rLvIA). High yields of fusion protein ranging from 100 to 500 mg/L culture were obtained. The pharmacological profile of rLvIA was determined by two-electrode voltage-clamp electrophysiology in Xenopus laevis oocytes expressing rat nAChR subtypes. The rLvIA antagonized the α3β2 nAChR subtype selectively with a nano-molar IC50. The rLvIA was analgesic in a mouse hot-plate test model of pain. Overall, this study provides an effective method to synthesize α-conotoxin LvIA in an E. coli recombinant expression system, and this approach could be useful to obtain active conopeptides in large quantity and at low cost. View Full-Text
Keywords: α-conotoxin LvIA; recombinant expression; fusion protein; nAChRs; electrophysiology; pain assay α-conotoxin LvIA; recombinant expression; fusion protein; nAChRs; electrophysiology; pain assay
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. (CC BY 4.0).

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MDPI and ACS Style

Zhu, X.; Bi, J.; Yu, J.; Li, X.; Zhang, Y.; Zhangsun, D.; Luo, S. Recombinant Expression and Characterization of α-Conotoxin LvIA in Escherichia coli. Mar. Drugs 2016, 14, 11.

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