Determinants of ion channel versus transporter mechanism in the K(+) transporter superfamily

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The constant traffic of substrates across membranes is controlled by two classes of proteins: channels and transporters. Open channels selectively allow a fast flux of substrates down an electrochemical gradient, whereas transporters are able to slowly pump substrates thermodynamically uphill by consuming energy. Due to these strictly opposing functions and the divergent speeds of substrate translocation, channels and transporters have traditionally been viewed as completely different entities. However, new structural and mechanistic data on both classes challenge this clear-cut separation. This proposal aims at elucidating the differences and commonalities of channels and transporters by studying the uniquely chimeric K(+) transport systems KtrAB and KdpFABC. In both systems, the translocating subunits KtrB and KpdA, respectively, are members of the superfamily of K(+) transporters, which are suggested to perform channel-like activities by themselves. However, in the presence of additional regulatory subunits both complexes are likely converted into transporters: The KdpFABC complex functions as a P-type ATPase while KtrAB probably exhibits Na(+)/K(+) symporter activity. By use of a multidisciplinary approach including transport studies, electrophysiology, EPR spectroscopy and X-ray crystallography, we will address the structure-function relationships underlying the mechanisms of K(+) translocation in both the isolated subunits KtrB and KdpA, respectively, and the assembled complexes. In particular, we will answer the question whether KtrAB in fact functions as symporter, how substrate translocation via the Ktr system is achieved and which structural changes in presence of the regulatory subunit KtrA cause the altered function. For the KdpFABC system we will focus on elucidating the mechanisms of how ATP hydrolysis in subunit KdpB is coupled to the active transport via KdpA and thus how structurally a former channel is converted into a primary active transporter. Taken together, this work will lead to basic understanding of what is requiered to make a transporter or channel.