{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,20]],"date-time":"2026-10-20T11:43:49Z","timestamp":1792496629972,"version":"4.5.0"},"reference-count":69,"publisher":"American Chemical Society (ACS)","issue":"20","license":[{"start":{"date-parts":[[2026,9,29]],"date-time":"2026-09-29T00:00:00Z","timestamp":1790640000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100000024","name":"Canadian Institutes of Health Research","doi-asserted-by":"publisher","award":["202209"],"award-info":[{"award-number":["202209"]}],"id":[{"id":"10.13039\/501100000024","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100003579","name":"University of Toronto","doi-asserted-by":"publisher","id":[{"id":"10.13039\/501100003579","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100000925","name":"National Health and Medical Research Council","doi-asserted-by":"publisher","award":["GNT2021638"],"award-info":[{"award-number":["GNT2021638"]}],"id":[{"id":"10.13039\/501100000925","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100000923","name":"Australian Research Council","doi-asserted-by":"publisher","award":["DP230102668"],"award-info":[{"award-number":["DP230102668"]}],"id":[{"id":"10.13039\/501100000923","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100001782","name":"University of Melbourne","doi-asserted-by":"publisher","id":[{"id":"10.13039\/501100001782","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2026,10,20]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>WalKR is a two-component system that regulates cell wall homeostasis and other processes in low-GC Gram-positive bacteria. It is essential for viability in Staphylococcus aureus and regulates genes that encode the autolysins and other critical proteins. The sensor kinase WalK phosphorylates WalR at the aspartic acid residue 53 (D53) in the receiver domain, stimulating promoter DNA binding. Unlike most response regulators, WalR is reported to have a second kinase, the serine\/threonine kinase PknB, which phosphorylates the receiver domain at the threonine residue 101. We previously reported that a walR mutation that changed T101 to a methionine conferred low-level resistance to vancomycin and greatly increased susceptibility to tunicamycin along with several other phenotypic traits. In this work, we demonstrate similarities between pknB null and the WalRT101M mutants. A combination of in vitro and in vivo data supports the idea that PknB interacts with WalR in living cells (consistent with similar experiments in Bacillus subtilis) and that this likely involves phosphorylation of WalR at T101. Using in silico modeling, we identified a possible intermolecular hydrogen bond between the T101 phosphate group and E108. This would have the effect of stabilizing the WalR dimerization interface. Consistent with this, we find that T101 is important for WalR dimer formation in vivo. The primary activation of WalR is clearly phosphorylation at D53 by WalK. However, our data suggest that PknB serves as a second potentiating input stabilizing the WalR dimer.<\/jats:p>","DOI":"10.1021\/acs.biochem.6c00334","type":"journal-article","created":{"date-parts":[[2026,9,29]],"date-time":"2026-09-29T19:02:06Z","timestamp":1790708526000},"page":"3167-3182","source":"Crossref","is-referenced-by-count":0,"title":["Phosphorylation\nby the PknB Serine\/Threonine Kinase\nStimulates Dimer Formation by WalR"],"prefix":"10.1021","volume":"65","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-9042-6659","authenticated-orcid":true,"given":"Marina","family":"Suppi","sequence":"first","affiliation":[{"name":"University of Toronto , , 661 University Avenue , , ,","place":["Toronto, Ontario, Canada, M5G 1M1"]},{"name":"Doherty Institute for Infection and Immunity, University of Melbourne , , , ,","place":["Melbourne, Victoria, Australia, 3000"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ian R.","family":"Monk","sequence":"additional","affiliation":[{"name":"Doherty Institute for Infection and Immunity, University of Melbourne , , , ,","place":["Melbourne, Victoria, Australia, 3000"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Liam K. R.","family":"Sharkey","sequence":"additional","affiliation":[{"name":"Doherty Institute for Infection and Immunity, University of Melbourne , , , ,","place":["Melbourne, Victoria, Australia, 3000"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2556-8316","authenticated-orcid":true,"given":"Nichollas E.","family":"Scott","sequence":"additional","affiliation":[{"name":"Doherty Institute for Infection and Immunity, University of Melbourne , , , ,","place":["Melbourne, Victoria, Australia, 3000"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dweipayan","family":"Goswami","sequence":"additional","affiliation":[{"name":"Doherty Institute for Infection and Immunity, University of Melbourne , , , ,","place":["Melbourne, Victoria, Australia, 3000"]},{"name":"University School of Sciences, Gujarat University , , ,","place":["Ahmedabad, Gujarat, India, 380009"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Aakash","family":"Natarajan","sequence":"additional","affiliation":[{"name":"University of Toronto , , 661 University Avenue , , ,","place":["Toronto, Ontario, Canada, M5G 1M1"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Stephanie","family":"Tan","sequence":"additional","affiliation":[{"name":"University of Toronto , , 661 University Avenue , , ,","place":["Toronto, Ontario, Canada, M5G 1M1"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Katharine","family":"Myler","sequence":"additional","affiliation":[{"name":"University of Toronto , , 661 University Avenue , , ,","place":["Toronto, Ontario, Canada, M5G 1M1"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sheila Marie","family":"Pimentel-Elardo","sequence":"additional","affiliation":[{"name":"University of Toronto , , 661 University Avenue , , ,","place":["Toronto, Ontario, Canada, M5G 1M1"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0150-123X","authenticated-orcid":true,"given":"Timothy P.","family":"Stinear","sequence":"additional","affiliation":[{"name":"Doherty Institute for Infection and Immunity, University of Melbourne , , , ,","place":["Melbourne, Victoria, Australia, 3000"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1202-6614","authenticated-orcid":true,"given":"Sacha J.","family":"Pidot","sequence":"additional","affiliation":[{"name":"Doherty Institute for Infection and Immunity, University of Melbourne , , , ,","place":["Melbourne, Victoria, Australia, 3000"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7909-1803","authenticated-orcid":true,"given":"Justin R.","family":"Nodwell","sequence":"additional","affiliation":[{"name":"University of Toronto , , 661 University Avenue , , ,","place":["Toronto, Ontario, Canada, M5G 1M1"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"316","published-online":{"date-parts":[[2026,9,29]]},"reference":[{"issue":"2","key":"2026102007164138100_cit1","first-page":"269","article-title":"Vancomycin Resistance in Staphylococcus aureus","volume":"90","author":"McGuinness","year":"2017","journal-title":"Yale J. 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Microbiol."},{"issue":"4","key":"2026102007164138100_cit4","doi-asserted-by":"publisher","first-page":"429.e1","DOI":"10.1016\/j.cmi.2017.07.028","article-title":"Emergence\nof Dalbavancin Non-Susceptible,\nVancomycin-Intermediate Staphylococcus aureus (VISA)\nafter Treatment of MRSA Central Line-Associated Bloodstream Infection\nwith a Dalbavancin- and Vancomycin-Containing Regimen","volume":"24","author":"Werth","year":"2018","journal-title":"Clin. Microbiol. Infect."},{"issue":"1","key":"2026102007164138100_cit5","doi-asserted-by":"publisher","DOI":"10.15252\/embr.202256184","article-title":"Current Developments\nin Antibiotic Discovery","volume":"24","author":"Hegemann","year":"2023","journal-title":"EMBO Rep."},{"issue":"6","key":"2026102007164138100_cit6","doi-asserted-by":"publisher","first-page":"3730","DOI":"10.1128\/AAC.00420-16","article-title":"Complete Reconstitution\nof the Vancomycin-Intermediate Staphylococcus aureus Phenotype of Strain Mu50 in Vancomycin-Susceptible S. aureus","volume":"60","author":"Katayama","year":"2016","journal-title":"Antimicrob. 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Dis."},{"issue":"8","key":"2026102007164138100_cit9","doi-asserted-by":"publisher","first-page":"3870","DOI":"10.1128\/AAC.01563-10","article-title":"walK and clpP Mutations Confer Reduced Vancomycin Susceptibility\nin Staphylococcus aureus","volume":"55","author":"Shoji","year":"2011","journal-title":"Antimicrob.\nAgents Chemother."},{"issue":"9","key":"2026102007164138100_cit10","doi-asserted-by":"publisher","first-page":"4188","DOI":"10.1128\/AAC.00398-11","article-title":"Mutation of RNA Polymerase \u03b2 Subunit (rpoB) Promotes hVISA-to-VISA Phenotypic Conversion of Strain\nMu3","volume":"55","author":"Matsuo","year":"2011","journal-title":"Antimicrob. 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Microbiol."},{"issue":"10","key":"2026102007164138100_cit16","doi-asserted-by":"publisher","first-page":"102473","DOI":"10.1016\/j.jbc.2022.102473","article-title":"A Defect\nin Cell Wall Recycling Confers Antibiotic\nResistance and Sensitivity in Staphylococcus aureus","volume":"298","author":"Tan","year":"2022","journal-title":"J. Biol. 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Immun."},{"issue":"22","key":"2026102007164138100_cit20","doi-asserted-by":"publisher","first-page":"8257","DOI":"10.1128\/JB.00645-07","article-title":"New Insights into the\nWalK\/WalR (YycG\/YycF) Essential Signal Transduction Pathway Reveal\na Major Role in Controlling Cell Wall Metabolism and Biofilm Formation\nin Staphylococcus aureus","volume":"189","author":"Dubrac","year":"2007","journal-title":"J.\nBacteriol."},{"issue":"1","key":"2026102007164138100_cit21","doi-asserted-by":"publisher","first-page":"523","DOI":"10.1038\/s41467-018-02949-y","article-title":"Sensory Deprivation\nin Staphylococcus aureus","volume":"9","author":"Villanueva","year":"2018","journal-title":"Nat.\nCommun."},{"issue":"6","key":"2026102007164138100_cit22","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pgen.1005275","article-title":"The Eukaryotic-Like\nSer\/Thr Kinase\nPrkC Regulates the Essential WalRK Two-Component System in Bacillus subtilis","volume":"11","author":"Libby","year":"2015","journal-title":"PLoS Genet."},{"issue":"1","key":"2026102007164138100_cit23","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1016\/j.ijmm.2016.12.001","article-title":"The Cell Wall Precursor Lipid II\nActs as a Molecular Signal for the Ser\/Thr Kinase PknB of Staphylococcus aureus","volume":"307","author":"Hardt","year":"2017","journal-title":"Int. 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Bacteriol."},{"issue":"4","key":"2026102007164138100_cit42","doi-asserted-by":"publisher","DOI":"10.1371\/journal.ppat.1004870","article-title":"Sigma Factor\nSigB Is Crucial to Mediate Staphylococcus aureus Adaptation\nduring Chronic Infections","volume":"11","author":"Tuchscherr","year":"2015","journal-title":"PLoS Pathog."},{"issue":"1","key":"2026102007164138100_cit43","doi-asserted-by":"publisher","first-page":"5133","DOI":"10.1038\/s41467-019-13127-z","article-title":"Multisite\nPhosphorylation Drives\nPhenotypic Variation in (p)ppGpp Synthetase-Dependent Antibiotic Tolerance","volume":"10","author":"Libby","year":"2019","journal-title":"Nat. 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