Study On Interaction Mechanism Of Celluar Membrane Expression Of Mslo Regulated By β2 In BK Channel

BK channel,Ca²⁺-activated voltage-dependent K⁺ channel plays a crucial role in modulating a number of important physiological processes,such as neuronal excitability, frequency tuning of hair cells,smooth muscle contraction,and immunity. Therefore, clarifying the structure and function of BK channel in molecular level is greatly vital for neuroscience research.In this study,we investigate the celluar membrane expression quantity of mslo(a homolog ofαsubunit),hβ2 (a homolog ofβsubunit)and its mutants influence on when hβ2 andαcoexpress in HEK293 cell with the technology of molecular biology,relevant immunology and patch-clamp. The main conclusions comprise as follows:Based on the work that the celluar membrane expression quantity of mslo influenced by hβ2 when hβ2 andαcoexpress in HEK293 cell is much larger than hβ2's mutant-dFIW-hβ2,we fuse dFIW(double FIW)(FIWFIW)artificially to different domains of hβ2 to validate that dFIW as a retain signal functions with special location and its detainment for celluar membrane expression quantity of mslo acts just when dFIW locates N terminal of hβ2.It is the hydrophobic amino acids in N terminal of hβ2 that affect the celluar membrane expression quantity of mslo when hβ2 andαcoexpress in HEK293 cell. The celluar membrane expression quantity of mslo is in proportion with the number of hydrophobic amino acids in N terminal of hβ2,that is to say,when hβ2 andαcoexpress, the stronger hydrophobic force of hβ2 N terminal,the less celluar membrane expression quantity of mslo. Moreover,the nearer N terminal hydrophobic amino acids locate,the greater influence on the celluar membrane expression quantity of mslo.A consistent conclusion concerns that BKβ2N-mediate pore occlusion exhibits shallow voltage dependence and is competed by the pore-blocking agent TEA,which puts forward a question of how far BKβ2N terminus enters the channel pore. In this study, we detect four-turn helix of BKβ2N terminus do not show an evident influence on regulating the celluar membrane expression quantity of mslo. Meanwhile,this conclusion further consolidates the importance of hydrophobic amino acids in BKβ2 N terminus.