Functional proteomics of BK potassium channels: defining the acuteoxygen sensor

摘要

Recombinant and native large conductance, Ca~(2+)-activated K~+ (BK) channels often demonstrate O_2 sensitivity in cell-free membrane patches suggesting that a significant component of the O_2-sensing machinery must be closely associated with the channel protein complex. Until recently, however, the identity of the O_2 sensor itself had remained elusive. Employing functional proteomics we have defined the molecular nature of such an O_2 sensor of BK channels. Using immunoprecipitation, ID and 2D gel electrophoresis, and mass spectroscopy we identified the constitutive form of haem oxygenase, haem oxygenase 2 (HO-2), as a BK alpha-subunit protein partner. Functional measurement of hypoxic modulation of BK channel activity during manipulation of HO-2 enzyme substrates and reaction products, followed by protein knock-down of HO-2 using small interfering RNA, indicated that this enzyme is direcdy involved in hypoxic inhibition of BK channels. Furthermore, good correlation was observed between data obtained from recombinant BK channels and those from acutely isolated rat carotid body glomus cells, suggesting strongly that HO-2 also acts as an O_2 sensor in native arterial chemoreceptors.