cbrA (PP_4695) encodes the sensor histidine kinase of the CbrA/CbrB two-component system, an atypical integral cytoplasmic-membrane protein in which an N-terminal solute/sodium symporter (SLC5) domain of about 13 transmembrane helices is joined through a STAC linker to cytoplasmic PAS, DHp and CA kinase domains. CbrA autophosphorylates on His766 and transfers the phosphoryl group to the receiver aspartate of its cognate response regulator CbrB; no CbrA phosphatase activity toward CbrB~P has been detected. The SLC5 domain specifically binds and takes up L-histidine, driven by the proton gradient, but L-histidine does not alter the kinase activities in vitro and the proximal signal sensed by CbrA remains unknown. Through CbrB, CbrA drives sigma-54-dependent expression of the CrcZ and CrcY small RNAs that antagonise Hfq/Crc translational repression, so the CbrA/CbrB system controls carbon catabolite repression and the balance of carbon and nitrogen utilization in pseudomonads and related Pseudomonadaceae.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0000155 phosphorelay sensor kinase activity | IEA GO_REF:0000002 | ACCEPT | Summary: Correct and core. Purified KT2440 CbrA autophosphorylates at the conserved His766 and transfers the phosphoryl group to CbrB, and the H766N variant fails to activate the CbrB target crcZ in vivo. Reason: The InterPro-based assignment is confirmed by direct biochemical evidence for KT2440 CbrA, including phosphotransfer to its cognate response regulator, which is the defining feature of a two-component sensor kinase. Supporting Evidence: PMID:32214184 it is shown that the histidine kinase can autophosphorylate and transfer the phosphoryl group to the response regulator CbrB. PMID:32214184 Contrary to wild type CbrA, CbrA-H766N did not activate expression of crcZ indicating that the conserved histidine in the DHp domain (site of phosphorylation) is essential for signal transduction. |
| GO:0000160 phosphorelay signal transduction system | IEA GO_REF:0000108 | ACCEPT | Summary: Correct and core. CbrA is the sensor component of the CbrA/CbrB two-component system. Reason: Both the KT2440 biochemistry and the original P. aeruginosa genetics define CbrA as the sensor kinase partner of CbrB in a His-Asp phosphorelay. Supporting Evidence: PMID:32214184 We show in in vitro experiments that CbrA autophosphorylates at the position of His766, and that the phosphoryl group is transferred to the response regulator CbrB. PMID:11401699 The cbrA and cbrB genes encode a sensor/histidine kinase (Mr 108 379, 983 residues) and a cognate response regulator (Mr 52 254, 478 residues) respectively. |
| GO:0004673 protein histidine kinase activity | IEA GO_REF:0000003 | MODIFY | Summary: Correct but less specific than the supported activity. CbrA is a two-component sensor kinase, which is the is_a child phosphorelay sensor kinase activity already carried by this gene. Reason: The EC-mapped term is true but general; replace with the more specific GO:0000155 phosphorelay sensor kinase activity, which this review accepts. Proposed replacements: phosphorelay sensor kinase activity Supporting Evidence: PMID:32214184 The DHp domain autophosphorylates under ATP consumption at His766, and transfers the phosphoryl group to Asp52 of CbrB. |
| GO:0006355 regulation of DNA-templated transcription | IEA GO_REF:0000002 | MARK AS OVER ANNOTATED | Summary: Over-annotation from the PAS-fold InterPro2GO mapping. CbrA does not bind DNA; it affects transcription only indirectly, by phosphorylating the transcription activator CbrB. Reason: A PAS domain does not imply transcription regulation, and the sensor kinase's own process is the phosphorelay. The transcriptional output (crcZ activation) is the activity of CbrB with sigma-54 and is annotated on those gene products. Supporting Evidence: PMID:32214184 CbrA is composed of a domain with similarity to proteins of the solute/sodium symporter family (SLC5) and domains typically found in bacterial sensor kinases. |
| GO:0007165 signal transduction | IEA GO_REF:0000120 | MODIFY | Summary: True but general; the specific process is the two-component phosphorelay already accepted for this gene. Reason: Replace with the is_a descendant GO:0000160 phosphorelay signal transduction system, which this review accepts. Proposed replacements: phosphorelay signal transduction system Supporting Evidence: PMID:32214184 We show in in vitro experiments that CbrA autophosphorylates at the position of His766, and that the phosphoryl group is transferred to the response regulator CbrB. |
| GO:0016020 membrane | IEA GO_REF:0000120 | MODIFY | Summary: Correct but general. CbrA is a polytopic protein of the cytoplasmic (inner) membrane, with an SLC5 transporter domain that takes up L-histidine into the cell and cytoplasmic kinase domains. Reason: Replace with GO:0005886 plasma membrane, the bacterial cytoplasmic membrane, consistent with the transporter topology and the uptake activity. Proposed replacements: plasma membrane Supporting Evidence: PMID:32214184 The SLC5 domain takes up L-histidine into the cell using an electrochemical proton gradient as driving force. PMID:11401699 The amino-terminal half (490 residues) of CbrA appears to be a sensor membrane domain, as predicted by 12 possible transmembrane helices file:PSEPK/cbrA/cbrA-uniprot.txt FT TRANSMEM 471..493 |
| GO:0016301 kinase activity | IEA GO_REF:0000104 | MODIFY | Summary: True but uninformative; the specific activity is phosphorelay sensor kinase activity. Reason: Replace with the is_a descendant GO:0000155 phosphorelay sensor kinase activity, which this review accepts. Proposed replacements: phosphorelay sensor kinase activity Supporting Evidence: PMID:32214184 it is shown that the histidine kinase can autophosphorylate and transfer the phosphoryl group to the response regulator CbrB. |
| GO:0016740 transferase activity | IEA GO_REF:0000104 | MODIFY | Summary: True but uninformative; the specific activity is phosphorelay sensor kinase activity. Reason: Replace with the is_a descendant GO:0000155 phosphorelay sensor kinase activity, which this review accepts. Proposed replacements: phosphorelay sensor kinase activity Supporting Evidence: PMID:32214184 it is shown that the histidine kinase can autophosphorylate and transfer the phosphoryl group to the response regulator CbrB. |
| GO:0016772 transferase activity, transferring phosphorus-containing groups | IEA GO_REF:0000002 | MODIFY | Summary: True but uninformative; the specific activity is phosphorelay sensor kinase activity. Reason: Replace with the is_a descendant GO:0000155 phosphorelay sensor kinase activity, which this review accepts. Proposed replacements: phosphorelay sensor kinase activity Supporting Evidence: PMID:32214184 it is shown that the histidine kinase can autophosphorylate and transfer the phosphoryl group to the response regulator CbrB. |
| GO:0022857 transmembrane transporter activity | IEA GO_REF:0000002 | MODIFY | Summary: Correct but general. The SLC5 domain of KT2440 CbrA is a specific, proton-gradient-driven L-histidine transporter, as shown by uptake assays in a strain lacking other histidine transporters. Reason: Replace with GO:0005290 L-histidine transmembrane transporter activity, an is_a descendant supported by direct transport and binding assays. Proposed replacements: L-histidine transmembrane transporter activity Supporting Evidence: PMID:32214184 It is demonstrated that CbrA can bind and transport L-histidine. Transport is specific for L-histidine and probably driven by an electrochemical proton gradient. PMID:26148710 Our combined data implicate the SSSF domain of CbrA in histidine transport and suggest that transport is coupled to signal transduction. |
| GO:0055085 transmembrane transport | IEA GO_REF:0000002 | MODIFY | Summary: Correct but general; the transported substrate is L-histidine. Reason: Replace with the is_a descendant GO:0089709 L-histidine transmembrane transport. Proposed replacements: L-histidine transmembrane transport Supporting Evidence: PMID:32214184 The results demonstrate that CbrA functions as a L-histidine transporter and sensor kinase. |
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Download this section (compressed HTML)Q: What intracellular or membrane-derived signal does CbrA sense, given that L-histidine binds CbrA but does not change its kinase activities in vitro?
Q: Is L-histidine uptake by the CbrA SLC5 domain physiologically significant in KT2440 relative to the dedicated HutT and HutWX transporters?
Experiment: Screen candidate intracellular metabolites (for example 2-oxoglutarate, glutamine, ATP/ADP ratio) for effects on CbrA-dependent phosphotransfer to CbrB in proteoliposomes, and correlate with crcZ reporter output in vivo.
Hypothesis: CbrA autokinase activity responds to an intracellular metabolite that tracks carbon/nitrogen balance.
Type: biochemical_assay
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