Quaternary Structure

The authors took advantage of sequence and structural similarities between the voltage- and calcium-activated BK channel homolog Slo1 [C. elegans] and the sodium-activated BK channel homolog Slo2 [in snails, birds, drosophila, frogs & mammals] to deduce that the quaternary structure of the human BK channel included a CTD tetramer.

An outline of the experimental approaches used:

1. Size exclusion chromatography on purified chicken Slo2.2 revealed that the CTD eluted as a tetramer. The stability of the complex was subsequently shown as it withstood the denaturing effects of SDS.
2. The Na⁺-activated Slo2 was crystallised in the presence of 500mM NaCl [ligand] which diffracted x-rays successfully with a resolution of 6Å.
3. Using a polyalanine model of BK CTD, a single solution in molecular replacement against the Slo2.2 diffraction data was obtained – which concluded that the CTD structures of the human BK and chicken Slo2.2 CTDs must have very similar structure. On application of crystallographic symmetry operators the tetrameric structure of the human BK CTD was elucidated at a high enough resolution to reveal the subunits but not high enough to extract atomic details. This allowed for the following model to be postulated:

Biological Evidence to support the tetrameric structure.

1. Corresponds well to the "gating ring" observed in the MthK chains

2. Amino acid pairs that are spatially far apart in the monomer have been shown to interact in double mutant cycles are close in proximity to each other through the creation of an assembly  interface.

However, there is a difference between the MthK and BK gating ring structures. The calcium binds on the assembly interface on the BK whereas it binds on the flexible interface on the MthK channel