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Structural and functional consequences of an amide-to-ester substitution in the selectivity filter of a potassium channel.

ABSTRACT: The selectivity filter of K(+) channels comprises four contiguous ion binding sites, S1 through S4. Structural and functional data indicate that the filter contains on average two K(+) ions at any given time and that these ions reside primarily in two configurations, namely to sites S1 and S3 or to sites S2 and S4. Maximum ion flux through the channel is expected to occur when the energy difference between these two binding configurations is zero. In this study, we have used protein semisynthesis to selectively perturb site 1 within the filter of the KcsA channel through use of an amide-to-ester substitution. The modification alters K(+) conduction properties. The structure of the selectivity filter is largely unperturbed by the modification, despite the loss of an ordered water molecule normally located just behind the filter. Introduction of the ester moiety was found to alter the distribution of K(+), Rb(+,) and Cs(+) within the filter, with the most dramatic change found for Rb(+). The redistribution of ions is associated with the appearance of a partially hydrated ion just external to the filter, at a position where no ion is observed in the wild-type channel. The appearance of this new ion-binding site creates a change in the distance between a pair of K(+) ions some fraction of the time, apparently leading to a reduction in the ion conduction rate. Importantly, this finding suggests that the selectivity filter of a potassium channel is optimized both in terms of absolute ion occupancy and in terms of the separation in distance between the conducting ions.

SUBMITTER: Valiyaveetil FI

PROVIDER: S-EPMC2546570 | biostudies-literature | 2006 Sep

REPOSITORIES: biostudies-literature

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Structural and functional consequences of an amide-to-ester substitution in the selectivity filter of a potassium channel.

Valiyaveetil Francis I FI Sekedat Matthew M MacKinnon Roderick R Muir Tom W TW

Journal of the American Chemical Society 20060901 35

The selectivity filter of K(+) channels comprises four contiguous ion binding sites, S1 through S4. Structural and functional data indicate that the filter contains on average two K(+) ions at any given time and that these ions reside primarily in two configurations, namely to sites S1 and S3 or to sites S2 and S4. Maximum ion flux through the channel is expected to occur when the energy difference between these two binding configurations is zero. In this study, we have used protein semisynthesi ...[more]

PMID: 16939283

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Project description:C-type inactivation is a time-dependent process of great physiological significance that is observed in a large class of K+ channels. Experimental and computational studies of the pH-activated KcsA channel show that the functional C-type inactivated state, for this channel, is associated with a structural constriction of the selectivity filter at the level of the central glycine residue in the signature sequence, TTV(G)YGD. The structural constriction is allosterically promoted by the wide opening of the intracellular activation gate. However, whether this is a universal mechanism for C-type inactivation has not been established with certainty because similar constricted structures have not been observed for other K+ channels. Seeking to ascertain the general plausibility of the constricted filter conformation, molecular dynamics simulations of a homology model of the pore domain of the voltage-gated potassium channel Shaker were performed. Simulations performed with an open intracellular gate spontaneously resulted in a stable constricted-like filter conformation, providing a plausible nonconductive state responsible for C-type inactivation in the Shaker channel. While there are broad similarities with the constricted structure of KcsA, the hypothetical constricted-like conformation of Shaker also displays some subtle differences. Interestingly, those are recapitulated by the Shaker-like E71V KcsA mutant, suggesting that the residue at this position along the pore helix plays a pivotal role in determining the C-type inactivation behavior. Free energy landscape calculations show that the conductive-to-constricted transition in Shaker is allosterically controlled by the degree of opening of the intracellular activation gate, as observed with the KcsA channel. The behavior of the classic inactivating W434F Shaker mutant is also characterized from a 10-μs MD simulation, revealing that the selectivity filter spontaneously adopts a nonconductive conformation that is constricted at the level of the second glycine in the signature sequence, TTVGY(G)D.

Dataset Information

Structural and functional consequences of an amide-to-ester substitution in the selectivity filter of a potassium channel.

Publications

Structural and functional consequences of an amide-to-ester substitution in the selectivity filter of a potassium channel.

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