Building 3-D Models of the Full-Length IKs Cnannel using Computational Techniques

Background: KCNQ1 (Q1, pore-forming) and KCNE1 (E1, regulatory) associate to form the IKs channel, a critical determinant of QT interval. None of Q1 homology models or Q1/E1 docking models includes the cytoplasmic domain (CD), despite its critical roles in IKs assembly/trafficking/modulation. One major obstacle is the very long (∼320 aa) and dynamic Q1 carboxyl terminus (CT). However, sub-regions of CT of Q1 and homologous KCNQ4 have been crystallized, and there is rich information on the functional roles of, and relationships among, sub-regions or residues in CD of Q1/E1. These prompt us to build 3-D models of full-length Q1/E1 (i.e. including CD) using a hierarchical approach.Methods: (1) Use Robetta server to predict structures of amino-terminus (NT, aa 1-140) and CT (aa 354-676) of Q1. (2) Select Robetta models compatible with existing structural data. (3) Remove flexible loop regions, and dock the helical regions of chosen Robetta models to the Q1 transmembrane homology model. This manual docking procedure is guided by data in the literature. (4) The most favored docking-configurations are triplicated to produce the full-length Q1 models. (5) Dock refined E1 NMR structure to the full-length Q1 model, guided by our disulfide-trapping data and information in the literature. (6) After energy-minimization and removing steric clashes, the systems will be subjected to molecular dynamics (MD) simulations. (7) Analyze the MD trajectories to design disulfide-trapping experiments for model validation or rejection.Results and Conclusion: We have narrowed down to three docking-configurations in step 4. Docking of E1 indicates that stoichiometries (E1:Q1) higher than 2:4 will make the system unstable, due to steric clashes between CT of Q1 and E1. Although we need to experimentally validate our models, our progress with computational approaches has provided insights into this critical region of IKs.