Nicotinic acetylcholine receptors (nAChRs) are a diverse family of homo-or heteropentameric ligand-gated ion channels. Understanding the physiological role of each nAChR subtype and the key residues responsible for normal and pathological states is important. α-Conotoxin neuropeptides are highly selective probes capable of discriminating different subtypes of nAChRs. In this study, we performed homology modeling to generate the neuronal α3, β2 and β4 subunits using the x-ray structure of the α1 subunit as a template. The structures of the extracellular domains containing ligand binding sites in the α3β2 and α3β4 nAChR subtypes were constructed using MD simulations and ligand docking processes in their free and ligand-bound states using α-conotoxin GIC, which exhibited the highest α3β2 vs. α3β4 discrimination ratio. The results provide a reasonable structural basis for such a discriminatory ability, supporting the idea that the present strategy can be used for future investigations on nAChR-ligand complexes.[BMB reports 2012; 45(5): 275-280]

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Mycobacterial cell-wall skeleton (CWS) is an immunoactive and biodegradable particulate adjuvant and has been tried to use for immunotherapy. The CWS of Mycobacterium bovis bacillus Calmette-Guerin (BCG-CWS) was studied as an universal vaccine vehicle for antigen conjugation, to develop potentially effective and safe vaccine. Although a variety of biological activities of BCG-CWS have been studied, the effects of BCG-CWS on spleen cells are not fully elucidated. Using MTT assay and trypan blue exclusion test, we found that BCG-CWS significantly enhanced the viability and proliferation of cells. Multiple clusters, indicating proliferation, were observed in BCG-CWS-treated spleen cells and surface marker staining assay revealed that BCG-CWS promoted the proliferation of CD19 + B lymphocyte rather than CD4 + or CD8 + T lymphocyte. In addition, BCG-CWS up-regulated the expression of anti-apoptotic molecules such as bcl-2, bcl-xL. BCG-CWS increased the surface expression of CD25 and CD69 as well as IL-2 production of spleen cells, suggesting increased activation. Furthermore, BCG-CWS enhanced the antigen-specific cell proliferation and interferon-gamma production of spleen cells. Taken together, these results demonstrate the immunostimulatory effects of BCG-CWS on spleen cells via multiple mechanisms, providing valuable information to broaden the use of BCG-CWS in clinical and research settings.

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