A Superfamily of Voltage-gated Sodium Channels in Bacteria

Abstract

NaChBac, a six-␣-helical transmembrane-spanning protein cloned from Bacillus halodurans, is the first functionally characterized bacterial voltage-gated Na ؉ -selective channel (Ren, D., Navarro, B., Xu, H., Yue, L., Shi, Q., and Clapham, D. E. (2001) Science 294, 2372-2375). As a highly expressing ion channel protein, NaChBac is an ideal candidate for high resolution structural determination and structure-function studies. The biological role of NaChBac, however, is still unknown. In this report, another 11 structurally related bacterial proteins are described. Two of these functionally expressed as voltage-dependent Na ؉ channels (Na V PZ from Paracoccus zeaxanthinifaciens and Na V SP from Silicibacter pomeroyi). Na V PZ and Na V SP share ϳ40% amino acid sequence identity with NaChBac. When expressed in mammalian cell lines, both Na V PZ and Na V SP were Na ؉ -selective and voltage-dependent. However, their kinetics and voltage dependence differ significantly. These single six-␣-helical transmembranespanning subunits constitute a widely distributed superfamily (Na V Bac) of channels in bacteria, implying a fundamental prokaryotic function. The degree of sequence homology (22-54%) is optimal for future comparisons of Na V Bac structure and function of similarity and dissimilarity among Na V Bac proteins. Thus, the Na V Bac superfamily is fertile ground for crystallographic, electrophysiological, and microbiological studies.Mammalian voltage-gated sodium (Na V ) 1 and calcium (Ca V ) channels underlie membrane excitability, muscle contraction, and hormone secretion (1). In contrast, the function of prokaryotic voltage-gated ion-selective channels is relatively unknown. Na ϩ channels may drive Na ϩ -dependent flagellar motors in certain marine and alkaliphilic species (2-6). In marine vibrio, PomAB and MotXY have been proposed to form a functional Na ϩ channel (5, 6), but the conductance has not been directly measured. In alkaliphilic bacteria, the prokaryotic ion channel responsible has not been identified. A bacterial 6-␣-helical transmembrane (6TM) channel subunit NaChBac was expressed in CHO cells as a functional voltage-gated Na ϩ channel (7), but its role in bacteria is still being elucidated.The pore-forming subunits (␣ 1 ) of mammalian Na V and Ca V are composed of four similar repeats of 6TM domains (8, 9), probably arising by gene duplication of a single 6TM gene (1, 10). The first bacterial voltage-gated Na ϩ channel (NaChBac) functionally expressed in mammalian cells was cloned from Bacillus halodurans (7). It contains a single 6TM domain of 274 amino acids but almost certainly forms a tetramer (7). NaChBac voltagedependent activation and inactivation kinetics are 10 -100 times slower than that of Na V (7). NaChBac inactivation may result from pore inactivation (C-type inactivation), since it does not contain an obvious cytoplasmic inactivation gate (7).Because single 6TM Na ϩ -selective ion channels do not appear to be present in vertebrates, the 24 TM structure of Na V s may have arisen under evolut...

Voltage-gated sodium channel (NaV) protein dissection creates a set of functional pore-only proteins

“…We previously identified an Na V from the marine sulfur-reducing bacterium S. pomeroyi (Na V Sp1), which is functional when expressed in mammalian cells (8), as a protein that could be highly expressed (approximately 20 mg L −1 ) in Escherichia coli membranes (15). Despite the high expression level, size-exclusion chromatography analysis of purified, detergent-solubilized Na V Sp1 revealed a broad elution profile indicative of a polydisperse sample that was not well suited to further characterization (Fig.…”

“…Functional studies of a number of bacterial Na V s have shown them to be sodium-selective (8)(9)(10)). Yet, in line with the close evolutionary relationship of Na V s and Ca V s (1-4), the bacterial Na V selectivity filter sequences also bear some sequence similarity to those of Ca V s (9,10,49).…”

“…Consequently, they are targets for a host of pharmaceuticals used to treat a diverse set of disorders and remain active targets for drug development (5)(6)(7). Recently, single subunit, six-transmembrane segment Na V s have been identified in a large number of bacteria from diverse environments (8,9). These channels show clear similarities to eukaryotic Na V s and Ca V s (2, 9, 10), suggesting that the prokaryotic channels may have been ancestors of the more complex vertebrate channels.…”

Astrocytes in Alzheimer's Disease

“…[2][3][4][5][6][7] The bacteria also evolved the first precursor of the Na ϩ channel, the NaChBac expressed, for example, in Bacillus halodurans. 8 In the eukaryotes, the excitable molecules developed further; in single-cell organisms, the waves of plasmalemmal excitation began to generate through the spreading opening/closures of voltagegated channels. 9 In parallel eukaryotic organisms acquired intracellular organelles, intracellular channels dwelling in the endomembranes, and exocytotic machinery formed the basis for chemical intercellular transmission.…”

Tetrameric Bacterial Sodium Channels: Characterization of Structure, Stability, and Drug Binding

“…Their function in mammals underlies neuronal signaling, muscle contraction, and hormone secretion (1). The first bacterial voltage-gated sodium channel (NaChBac) 1 was identified from Bacillus halodurans (2), and a number of orthologs were subsequently identified, forming a superfamily (3). The function of voltage-gated sodium channels in bacteria is still a matter of debate; however, there is evidence that these channels are involved in processes such as motility, chemotaxis, and pH homeostasis (4).…”