Key Residues in the Nicotinic Acetylcholine Receptor β2 Subunit Contribute to α-Conotoxin LvIA Binding |
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Authors: | Dongting Zhangsun Xiaopeng Zhu Yong Wu Yuanyan Hu Quentin Kaas David J Craik J Michael McIntosh Sulan Luo |
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Institution: | From the ‡Key Laboratory of Tropical Biological Resources, Ministry of Education, Key Lab for Marine Drugs of Haikou, Hainan University, Haikou, Hainan 570228, China.;the §Institute for Molecular Bioscience, The University of Queensland, Brisbane, Queensland 4072 Australia, and ;the ¶George E. Wahlen Veterans Affairs Medical Center and Departments of Biology and Psychiatry, University of Utah, Salt Lake City, Utah 84112 |
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Abstract: | α-Conotoxin LvIA (α-CTx LvIA) is a small peptide from the venom of the carnivorous marine gastropod Conus lividus and is the most selective inhibitor of α3β2 nicotinic acetylcholine receptors (nAChRs) known to date. It can distinguish the α3β2 nAChR subtype from the α6β2* (* indicates the other subunit) and α3β4 nAChR subtypes. In this study, we performed mutational studies to assess the influence of residues of the β2 subunit versus those of the β4 subunit on the binding of α-CTx LvIA. Although two β2 mutations, α3β2F119Q] and α3β2T59K], strongly enhanced the affinity of LvIA, the β2 mutation α3β2V111I] substantially reduced the binding of LvIA. Increased activity of LvIA was also observed when the β2-T59L mutant was combined with the α3 subunit. There were no significant difference in inhibition of α3β2T59I], α3β2Q34A], and α3β2K79A] nAChRs when compared with wild-type α3β2 nAChR. α-CTx LvIA displayed slower off-rate kinetics at α3β2F119Q] and α3β2T59K] than at the wild-type receptor, with the latter mutant having the most pronounced effect. Taken together, these data provide evidence that the β2 subunit contributes to α-CTx LvIA binding and selectivity. The results demonstrate that Val111 is critical and facilitates LvIA binding; this position has not previously been identified as important to binding of other 4/7 framework α-conotoxins. Thr59 and Phe119 of the β2 subunit appear to interfere with LvIA binding, and their replacement by the corresponding residues of the β4 subunit leads to increased affinity. |
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Keywords: | Docking Receptor Receptor Structure-Function Receptor-interacting Protein (RIP) Toxin β subunit contribution α -Conotoxin LvIA α 3β 2 nAChR Mutant α 3β 2 Subtype |
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