DVU_1606 (Q72BM9) is a TrkA-family potassium uptake regulatory protein containing RCK_N and RCK_C domains. Based on structural homology to characterized KtrA/TrkA proteins from other bacteria, it functions as the cytosolic regulatory subunit of a Trk/Ktr K+ uptake system. The protein assembles into an octameric RCK gating ring that associates with a membrane-embedded TrkH/KtrB pore. ATP binding promotes an active square-like conformation that opens the pore for K+ flux, while ADP binding induces an inactive diamond-like conformation. Na+ binding at intra-dimer sites synergistically stabilizes the active state. The protein may also bind c-di-AMP, which typically reduces K+ import by destabilizing regulator-pore interactions. The core physiological role is regulation of K+ homeostasis and osmoadaptation.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0098655 monoatomic cation transmembrane transport | IEA GO_REF:0000108 | MODIFY | Summary: This annotation was inferred from GO:0008324 (transporter activity) via logical inference. However, Q72BM9 is NOT the membrane transporter itself but the cytosolic regulatory RCK subunit that gates the TrkH/KtrB pore. While the protein is involved in regulating cation transport, it does not itself participate in the transmembrane transport process. The annotation should be modified to reflect regulatory involvement rather than direct transport. Reason: TrkA/KtrA family proteins are cytosolic regulatory subunits that form octameric RCK gating rings to modulate the activity of membrane K+ pores (TrkH/KtrB). Cryo-EM structures of B. subtilis KtrAB at 2.8 A resolution demonstrate that KtrA forms an octameric ring bound to dimeric KtrB, with ATP/ADP binding controlling pore gating rather than direct transport (Chiang et al. 2024). The protein regulates K+ transmembrane transport but does not itself transport ions across the membrane. Proposed replacements: regulation of potassium ion transmembrane transport Supporting Evidence: file:DESVH/Q72BM9/Q72BM9-deep-research-falcon.md DVU_1606 (TrkA-family RCK protein) is a cytosolic/peripheral protein that assembles into an octameric RCK gating ring associated with an inner-membrane K+ pore DOI:10.1038/s41467-024-48057-y The regulatory subunits, TrkA and KtrA, which are referred to as reg- ulator of K+ conductance (RCK) proteins, can bind to signaling mole- cules in the cytosol and control the gating of the transmembrane pores, TrkH and KtrB, respectively. |
| GO:0005886 plasma membrane | IEA GO_REF:0000044 | MODIFY | Summary: This annotation derives from UniProtKB subcellular location vocabulary mapping, which incorrectly predicts Q72BM9 as a multi-pass membrane protein based on Phobius transmembrane helix predictions. However, characterized TrkA/KtrA homologs are cytosolic proteins that associate with the membrane pore peripherally via protein-protein interactions, not as integral membrane proteins. The RCK domains (positions 127-350) are clearly cytosolic based on structural data from homologs. Reason: Structural biology of KtrA/TrkA proteins demonstrates they are cytosolic regulatory subunits. The 2024 cryo-EM structure of B. subtilis KtrAB shows KtrA as a cytosolic octameric ring that binds to the cytoplasmic face of the KtrB membrane pore. The RCK_N and RCK_C domains that comprise the functional core of this protein are well-established as cytosolic ligand-binding domains. While Phobius predicts two N-terminal transmembrane helices, the dominant functional domains are cytosolic, and the protein should be annotated to cytoplasm with peripheral association to the plasma membrane. Proposed replacements: cytoplasm Supporting Evidence: DOI:10.1038/s41467-024-48057-y Both systems share a common protein quaternary structure: transmembrane subunits responsible for K+ permeation, and regulatory subunits located in the cytosol, forming a ring-like structure for association with the transmembrane subunits. |
| GO:0006813 potassium ion transport | IEA GO_REF:0000002 | ACCEPT | Summary: This annotation from InterPro (IPR003148, IPR006037, IPR036721) captures the involvement of Q72BM9 in potassium ion transport. While the protein does not directly transport K+, it is an essential regulatory component of the K+ uptake system. The term GO:0006813 is appropriately general (covering both direct transport and regulatory involvement) and is acceptable, though a more specific regulatory term would be more informative. Reason: GO:0006813 (potassium ion transport) describes the directed movement of K+ ions by means of transporters or pores, which includes regulatory subunits that are essential for this process. TrkA/KtrA proteins are obligate components of the functional Trk/Ktr K+ uptake system, and without them the pore cannot be properly gated. The annotation is acceptable as it captures the biological process in which the protein participates, even though its specific role is regulatory rather than catalytic. The two-component architecture on which this rests - a membrane K+ permeation subunit (TrkH/KtrB) plus a cytosolic RCK regulatory ring (TrkA/KtrA) - is documented by Chiang et al. 2024 (DOI:10.1038/s41467-024-48057-y); the c-di-AMP review of Foster et al. 2024 (DOI:10.1128/mmbr.00181-23) is cached here as summary text only, so no verbatim passage from it is quoted. |
| GO:0008324 monoatomic cation transmembrane transporter activity | IEA GO_REF:0000002 | MODIFY | Summary: This MF annotation from InterPro (IPR006037 - RCK_C) is an over-annotation. Q72BM9 is the regulatory RCK subunit, NOT the membrane transporter itself. RCK domains bind nucleotides (ATP/ADP) and undergo conformational changes that regulate the associated membrane pore, but they do not themselves enable transmembrane transport. The appropriate MF annotation would be potassium channel regulator activity or transporter regulator activity. Reason: Transporter activity terms should be reserved for proteins that directly enable the movement of substrates across membranes. TrkA/KtrA RCK proteins regulate transporter activity by binding to and modulating the TrkH/KtrB membrane pore. The 2024 structural study demonstrates that KtrA binds ATP/ADP at RCK interfaces and undergoes square-to-diamond conformational transitions that mechanically gate the associated pore. This is regulator activity, not transporter activity. The most appropriate term is GO:0015459 (potassium channel regulator activity) which specifically describes proteins that bind to and modulate K+ channels/transporters. Proposed replacements: potassium channel regulator activity ATP binding Supporting Evidence: DOI:10.1038/s41467-024-48057-y ATP binding to KtrA octamer activates K + ο¬ux activity of KtrB, whereas ADP binding inactivates it DOI:10.1099/mic.0.001597 In Streptococcus mitis, TrkA directly binds c-di-AMP and is required for growth under low K+. Loss of TrkA compromises growth and modulates cellular c-di-AMP levels, reinforcing TrkA's role in c-di-AMP-mediated K+ control. |
| GO:0015459 potassium channel regulator activity | ISS DOI:10.1038/s41467-024-48057-y | NEW | Summary: NEW annotation. Based on structural and functional characterization of homologous KtrA proteins, Q72BM9 functions as a potassium channel/transporter regulator. It binds to and modulates the activity of the associated TrkH/KtrB K+ pore through nucleotide-dependent conformational changes. Reason: The core molecular function of TrkA/KtrA proteins is to regulate the activity of associated K+ transporters. GO:0015459 (potassium channel regulator activity) accurately captures this function. Evidence from structural studies of B. subtilis KtrAB demonstrates that KtrA binds to KtrB and modulates its activity through ATP/ADP-dependent conformational changes. Supporting Evidence: DOI:10.1038/s41467-024-48057-y ATP binding to KtrA octamer activates K + ο¬ux activity of KtrB, whereas ADP binding inactivates it |
| GO:0005524 ATP binding | ISS DOI:10.1038/s41467-024-48057-y | NEW | Summary: NEW annotation. The RCK_N domain of TrkA/KtrA proteins contains ATP/ADP binding sites that are essential for the regulatory function. ATP binding at intra-dimer interfaces stabilizes the active conformation. Reason: ATP binding is a core biochemical function of TrkA/KtrA RCK proteins. The NAD(P)-binding domain superfamily signature (IPR036291) and structural studies confirm nucleotide binding capability. ATP/ADP binding is the primary mechanism by which these proteins sense cellular energy status and regulate K+ uptake accordingly. Supporting Evidence: DOI:10.1038/s41467-024-48057-y Altogether, the results suggest that Na+ binding at the BsKtrA intra-dimer interface coordinated by theΞ³-phosphates of ATPs and the carboxylate groups of Glu125 stabilizes the square-shaped ATP-BsKtrA octameric ring and maintains the pore-open conformation of BsKtrB |
| GO:0180001 cyclic-di-AMP binding | ISS DOI:10.1128/mmbr.00181-23 | NEW | Summary: NEW annotation. TrkA/KtrA proteins are known receptors for c-di-AMP, a bacterial second messenger that regulates K+ homeostasis. c-di-AMP binding to RCK domains typically reduces K+ import by destabilizing regulator-pore interactions. Reason: c-di-AMP is a master regulator of K+ homeostasis in bacteria, and TrkA/KtrA RCK proteins are established c-di-AMP receptors. Direct binding has been demonstrated for TrkA in S. mitis (Vevik 2025). The reported binding affinities across different RCK/CBS-containing K+ transport regulators range from ~40 nM to low uM; that range is taken from the Foster et al. 2024 review (DOI:10.1128/mmbr.00181-23), which is cached here as summary text only, so it is asserted here rather than quoted. This is a likely function for Q72BM9 based on conservation of the RCK domain architecture. Supporting Evidence: DOI:10.1099/mic.0.001597 In Streptococcus mitis, TrkA directly binds c-di-AMP and is required for growth under low K+. |
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Download this section (compressed HTML)Q: What is the cognate TrkH/KtrB membrane pore partner for DVU_1606 in D. vulgaris?
Q: Does DVU_1606 bind c-di-AMP, and if so, with what affinity?
Q: Is there evidence for Na+ synergy in DVU_1606 function as seen in B. subtilis KtrA?
Experiment: Isothermal titration calorimetry to measure ATP, ADP, and c-di-AMP binding affinities
Hypothesis: DVU_1606 binds ATP with higher affinity than ADP, and also binds c-di-AMP
Experiment: Co-immunoprecipitation or bacterial two-hybrid to identify the cognate membrane pore partner
Hypothesis: DVU_1606 interacts with a TrkH/KtrB homolog in D. vulgaris
Experiment: Cryo-EM structure of DVU_1606 octameric ring to confirm conservation of gating mechanism
Hypothesis: DVU_1606 forms an octameric ring similar to B. subtilis KtrA
Experiment: K+ uptake assays in D. vulgaris DVU_1606 deletion mutant under various osmotic conditions
Hypothesis: DVU_1606 is required for efficient K+ uptake and osmoadaptation
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