GPMDB002544OtherCorradini E, et al.InstrumentNot AvailableNot availableHomo_sapiens_viruses, Human_femalea.j.r.heck@uu.nl25653285http://gpmdb.thegpm.org/~/dblist_gpmnum/gpmnum=GPM32320007636http://gpmdb.thegpm.org/~/dblist_gpmnum/gpmnum=GPM32320007632http://gpmdb.thegpm.org/~/dblist_gpmnum/gpmnum=GPM32320007633http://gpmdb.thegpm.org/~/dblist_gpmnum/gpmnum=GPM32320007634http://gpmdb.thegpm.org/~/dblist_gpmnum/gpmnum=GPM32320007631Utrecht University, NetherlandsNot availableGPMDBProtein-protein interactions are important in providing compartmentalization and specificity in cellular signal transduction. Many studies have hallmarked the well designed compartmentalization of the cAMP-dependent protein kinase (PKA) through its anchoring proteins. Much less data are available on the compartmentalization of its closest homolog, cGMP-dependent protein kinase (PKG), via its own PKG anchoring proteins (GKAPs). For the enrichment, screening, and discovery of (novel) PKA anchoring proteins, a plethora of methodologies is available, including our previously described chemical proteomics approach based on immobilized cAMP or cGMP. Although this method was demonstrated to be effective, each immobilized cyclic nucleotide did not discriminate in the enrichment for either PKA or PKG and their secondary interactors. Hence, with PKG signaling components being less abundant in most tissues, it turned out to be challenging to enrich and identify GKAPs. Here we extend this cAMP-based chemical proteomics approach using competitive concentrations of free cyclic nucleotides to isolate each kinase and its secondary interactors. Using this approach, we identified Huntingtin-associated protein 1 (HAP1) as a putative novel GKAP. Through sequence alignment with known GKAPs and secondary structure prediction analysis, we defined a small sequence domain mediating the interaction with PKG Iβ but not PKG Iα. In vitro binding studies and site-directed mutagenesis further confirmed the specificity and affinity of HAP1 binding to the PKG Iβ N terminus. These data fully support that HAP1 is a GKAP, anchoring specifically to the cGMP-dependent protein kinase isoform Iβ, and provide further evidence that also PKG spatiotemporal signaling is largely controlled by anchoring proteins.Huntingtin-associated protein 1 (HAP1) is a cGMP-dependent kinase anchoring protein (GKAP) specific for the cGMP-dependent protein kinase Iβ isoform.Corradini Eleonora E,Burgers Pepijn P PP,Plank Michael M,Heck Albert J R AJ,Scholten Arjen A,Corradini Eleonora E, Burgers Pepijn P PP, Plank Michael M, Heck Albert J R AJ, Scholten Arjen ADLGAP1B, DAP1, DLGAP1A, APEN, HIP5, APX, DAP-1, anchoring, cGKI, GKPA|SAPAP, Gkap, BB075781, hHLP1, GKAP|SAPAP, Phosphotransferases, hGKAP, polypeptide, cGK1, AI845682, DAP-1-ALPHA, D17Bwg0511e, GKAP, Kinase, REF1, HAP2, HAP1, SAPAP1, APEX, mKIAA4162, 4933422O14Rik, 9630002F18, Transphosphorylases, cGMP-dependent protein kinase I., AI848168, DAP-1-BETA, Kinases, cGK 1, HLP, Sapap1, HAP-1, proteins, HAP1-B, HAP1-A, Apex, Ref-1, APE1, 2.7.11.12, APE, associated, ATP Phosphotransferases, ATPDLGAP1B, DAP1, DLGAP1A, CRAMP, APEN, APX, conformation, determination, FALL39, GKPA|SAPAP, rad/s^[2], Gkap, Alignment, ATP:protein phosphotransferase (cGMP-dependent) activity, 11H2, 10, (H, 11, 12, 17, AI845682, cGKI-alpha, reduced, Cyclic adenylic acid, D17Bwg0511e, 18)21-7/h2-4, CAP-18, 1, GKAP, 3, 18)(H2, Kinase, tiny, REF1, C13orf8, APEX, mKIAA4162, 9630002F18, 4aR, PRKGR1B, DAP-1-BETA, Determination, cAMP, 7-diol 2-oxide, 3':5'-cyclic GMP-dependent protein kinase activity, cGK 1, HAP-1, InChIKey=IVOMOUWHDPKRLL-BJEHYBLCDS, Sequence Homology Determination, hypoplasia, ZNF828, 13)/t4-, proteins, 7aS)-6-(6-amino-9H-purin-9-yl)tetrahydro-4H-furo[3, HAP1-B, HAP1-A, free, Apex, 6-7, Concentration, Concentrations, Cyclic Nucleotides, STK23, cGMP-dependent protein kinase ibeta activity, Nc1ncnc2n(cnc12)[C@@H]1O[C@@H]2COP(O)(=O)O[C@H]2[C@H]1O, Alignments, associated, ATP Phosphotransferases, relational structural quality, ATP, Sequence Homology Determinations, small, data, InChI=1/C10H12N5O6P/c11-8-5-9(13-2-12-8)15(3-14-5)10-6(16)7-4(20-10)1-19-22(17, cGK, HIP5, (2R, DAP-1, cGKI, BB075781, hHLP1, CAP18, AAT8, GKAP|SAPAP, 5'-phosphate, Phosphotransferases, PKG 1beta, CAMP, Determinations, PKG 1alpha, hGKAP, polypeptide, Sequence Alignments, cGK1, PRKG1B, DAP-1-ALPHA, Cyclic AMP, Sequence, PKG, chemical analysis, sequence, cGKI-BETA, 6-, HAP2, HAP1, SAPAP1, 2]dioxaphosphinine-2, 4933422O14Rik, 16H, Transphosphorylases, AI848168, LL37, FALL-39, mating_type_alpha, underdeveloped, Kinases, HLP, Sapap1, CHAMP, Sequence Homology, 2-d][1, PKG II, 5'-(hydrogen phosphate), 7-, alpha, 10-/m1/s1/f/h17H, 1H2, 5'-CYCLIC-MONOPHOSPHATE, 6R, primary structure of sequence macromolecule, C10H12N5O6P, metastatic, Ref-1, APE1, alpha mating type (yeast)., guanosine 3':5'-cyclic monophosphate-dependent protein kinase activity, Adenosine 3', ADENOSINE-3', assay, APE, adenosine 3', 7R, AttentionsDLGAP1B, DAP1, DLGAP1A, APEN, HIP5, APX, DAP-1, anchoring, cGKI, GKPA|SAPAP, Gkap, BB075781, hHLP1, GKAP|SAPAP, Phosphotransferases, hGKAP, polypeptide, cGK1, AI845682, DAP-1-ALPHA, D17Bwg0511e, GKAP, Kinase, REF1, HAP2, HAP1, SAPAP1, APEX, mKIAA4162, 4933422O14Rik, 9630002F18, Transphosphorylases, cGMP-dependent protein kinase I., AI848168, DAP-1-BETA, Kinases, cGK 1, HLP, Sapap1, HAP-1, proteins, HAP1-B, HAP1-A, Apex, Ref-1, APE1, 2.7.11.12, APE, associated, ATP Phosphotransferases, ATPSignal Transductions, protein kinase A activity, APX, determination, FALL39, Gkap, 10, 11, 12, Receptor Mediated Signal Transduction, 17, cGKI-alpha, AI845682, Method, cAMP-dependent protein kinase, 18)21-7/h2-4, CAP-18, 1, GKAP, 3, 18)(H2, Kinase, REF1, C13orf8, mKIAA4162, 9630002F18, PRKGR1B, DAP-1-BETA, reference sample, Determination, cAMP, 7-diol 2-oxide, HAP-1, Tissue, InChIKey=IVOMOUWHDPKRLL-BJEHYBLCDS, Sequence Homology Determination, hypoplasia, ZNF828, Pathways, 3', 5' cAMP-dependent protein kinase complex, proteins, free, 6-7, 5' cAMP-dependent protein kinase activity, Methodological Studies, signaling process, Concentrations, Signal Transduction Pathways, Transductions, Nc1ncnc2n(cnc12)[C@@H]1O[C@@H]2COP(O)(=O)O[C@H]2[C@H]1O, Alignments, associated, ATP Phosphotransferases, ATP, single organism signaling, Sequence Homology Determinations, Pathway, cGK, HIP5, (2R, anchoring, cGKI, cyclic AMP-dependent protein kinase complex, CAP18, Procedure, AAT8, GKAP|SAPAP, 5'-phosphate, CAMP, Determinations, PKG 1alpha, signaling cascade, cGK1, PRKG1B, Cyclic AMP, 5'-cAMP-dependent protein kinase complex, cGKI-BETA, 6-, Transduction, HAP2, HAP1, 4933422O14Rik, 16H, Transphosphorylases, PKA C, AI848168, System, CHAMP, Sequence Homology, PKG II, 7-, Methodological, 1H2, Methodological Study, 6R, C10H12N5O6P, metastatic, APE1, ATP:protein phosphotransferase (cAMP-dependent) activity, signaling pathway, guanosine 3':5'-cyclic monophosphate-dependent protein kinase activity, Adenosine 3', adenosine 3', 7R, DLGAP1B, DAP1, DLGAP1A, Signal Transduction Systems, CRAMP, biological signaling, APEN, Procedures, conformation, GKPA|SAPAP, Receptor-Mediated, Alignment, Gene, Signal Transduction System, ATP:protein phosphotransferase (cGMP-dependent) activity, 11H2, (H, Signal Transduction, method, reduced, Cyclic adenylic acid, method used in an experiment, D17Bwg0511e, Gene Products, Studies, tiny, AMPK, APEX, 4aR, 3':5'-cyclic GMP-dependent protein kinase activity, cGK 1, ligand, Signal, 13)/t4-, Signal Pathways, 7aS)-6-(6-amino-9H-purin-9-yl)tetrahydro-4H-furo[3, HAP1-B, Signal Transduction Pathway, HAP1-A, Apex, Study, signalling pathway, 5'-cyclophosphate-dependent protein kinase activity, Concentration, Cyclic Nucleotides, STK22, STK23, cGMP-dependent protein kinase ibeta activity, Signal Pathway, relational structural quality, Controlled, small, Gene., InChI=1/C10H12N5O6P/c11-8-5-9(13-2-12-8)15(3-14-5)10-6(16)7-4(20-10)1-19-22(17, data, Controlling, 5'-cyclophosphate-dependent protein kinase complex, DAP-1, Proteins, Receptor-Mediated Signal Transductions, BB075781, hHLP1, Phosphotransferases, PKG 1beta, hGKAP, polypeptide, Sequence Alignments, cyclic AMP-dependent protein kinase activity, PKA, DAP-1-ALPHA, Sequence, Receptor-Mediated Signal Transduction, PKG, Systems, Protein, chemical analysis, Mutageneses, sequence, signalling cascade, intrinsic catalyst activity, SAPAP1, 2]dioxaphosphinine-2, LL37, FALL-39, underdeveloped, Kinases, HLP, Sapap1, 2-d][1, 5'-(hydrogen phosphate), 5'-cAMP-dependent protein kinase activity, 10-/m1/s1/f/h17H, 5'-CYCLIC-MONOPHOSPHATE, primary structure of sequence macromolecule, signalling, Protein Gene Products, plan specification, Gene Proteins, Ref-1, signalling process, ADENOSINE-3', assay, APE, Attentions25044http://gpmdb.thegpm.org/~/dblist_gpmnum/gpmnum=GPM32320007631dbgap_ncbi~0patentfamilies~0rfam~0merops~0complex-portal~0uniprot~0wormbaseparasite~0embl-covid19~0reactome~0emdb~0wgs_masters~0ebiweb_resources~0opentargets_genetics~0biomodels_all~0intact~39ipd-mhc~0ebiweb_teams~0taxonomy~0genome_assembly~0sc-experiments~0ebiweb_people~0enzymeportal_enzymes~0ipd-nhkir~0cellosaurus~0pdbe~0chebi~0patentproteins~0interpro7~0uniref~0chembl~0pdbekb~0gpcrdb~0hgnc~0sc-genes~0rhea~0ebiweb_training~0alphafold~0imgt-hla~0patentnucleotides~0ensemblroot~0eva_studies~0non-coding~0europepmc~0pubmed~1identifiers_registry~0pdbechem~0hpa-covid19~0eva-variants-covid19~0biosamples~0gwas_catalog~0biotools~0tls_masters~0mesh~0coding~0sra~0opentargets~0efo~0embl-pathogen~0project~0pride~1human_diseases~0geo_datasets~0embl~0treefam~0uniparc~0ols~0dgva~0intenz~0go~0tsa_masters~0biosamples-covid19~0ebiweb_corporate~0omim~0lrg~0earlycause-molecular-sequences~0ipd-kir~0empiar~0rnacentral~0orcid_data_claims~0gpmdb~2lineage-covid19~0metagenomics~0pfam~0pride archive~1varsite~00Resource Reanalysis0.00.00.00771128932757557051.00.0falseHuntingtin associated protein (HAP1) is a cGMP-dependent kinase anchoring protein (GKAP), specific for the cGMP-dependent protein kinase Iβ isoform.Data from ProteomeXchange, PXD ID: PXD001434. Published as part of J Biol Chem. 2015 Feb 4. pii . From the Abstract: {{i}} ... Here, we extend on this cAMP-based chemical proteomics approach, employing competitive concentrations of free cyclic nucleotides to isolate each kinase and its secondary interactors. Using this approach we identified Huntingtin associated protein 1 (HAP1) as a putative novel GKAP. Through sequence alignment with known GKAPs and secondary structure prediction analysis we defined a small sequence domain mediating the interaction with PKG Iβ, but not PKG I alpha ... {{/i}}2015-02-10GPM3232000763125653285PXD001434PXD001434