{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,10,12]],"date-time":"2026-10-12T04:26:30Z","timestamp":1791779190049,"version":"4.3.4"},"posted":{"date-parts":[[2026,1,9]]},"group-title":"Microbiology","reference-count":57,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2026,1,9]],"date-time":"2026-01-09T00:00:00Z","timestamp":1767916800000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"funder":[{"name":"NIH","award":["R21AI182950"],"award-info":[{"award-number":["R21AI182950"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2026,1,12]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>\n                  <jats:italic>Staphylococcus aureus<\/jats:italic>\n                  uses the adhesin clumping factor A (ClfA) to bind fibrinogen and promote invasive infection through two distinct interfaces: an exposed, low-affinity site on the N3 head domain and a buried, high-affinity \u201cdock, lock, and latch\u201d (DLL) trench that is exposed only under shear. This dual-interface architecture allows limited antibody penetration, as antibodies typically block only the exposed site. Here, we establish a dual-mode inhibition strategy that overcomes this constraint by combining a high-affinity ClfA-binding designed ankyrin repeat protein (DARPin) with a fibrinogen v-chain peptide capable of occupying the DLL trench. Using cell-free click display and kinetics-guided affinity maturation, we engineer DARPin\u2013v-peptide fusion biologics that simultaneously block both fibrinogen-binding interfaces. These molecules inhibit ClfA\u2013fibrinogen interactions, prevent methicillin-resistant\n                  <jats:italic>S. aureus<\/jats:italic>\n                  agglutination in human plasma, neutralize major clinical ClfA variants, and confer Fc-independent protection in a lethal murine bacteremia model. This work provides a strategy for targeting antibody-intractable force-activated staphylococcal adhesins.\n                <\/jats:p>","DOI":"10.64898\/2026.01.09.698026","type":"posted-content","created":{"date-parts":[[2026,1,10]],"date-time":"2026-01-10T03:05:15Z","timestamp":1768014315000},"source":"Crossref","is-referenced-by-count":1,"title":["Dual-mode ClfA-targeting DARPin biologics protect against diverse methicillin-resistant\n                  <i>Staphylococcus aureus<\/i>\n                  strains"],"prefix":"10.64898","author":[{"given":"Karuppiah","family":"Chockalingam","sequence":"first","affiliation":[{"name":"Department of Microbial Pathogenesis and Immunology, Texas A&M College of Medicine, Bryan, TX, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Biswarup","family":"Banerjee","sequence":"additional","affiliation":[{"name":"Department of Microbiology, Howard Taylor Ricketts Laboratory, The University of Chicago, Lemont, IL, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yu","family":"Zeng","sequence":"additional","affiliation":[{"name":"Department of Microbial Pathogenesis and Immunology, Texas A&M College of Medicine, Bryan, TX, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shuyu","family":"Miao","sequence":"additional","affiliation":[{"name":"Department of Microbial Pathogenesis and Immunology, Texas A&M College of Medicine, Bryan, TX, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Joshua","family":"Bryant","sequence":"additional","affiliation":[{"name":"Department of Microbial Pathogenesis and Immunology, Texas A&M College of Medicine, Bryan, TX, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dominique","family":"Missiakas","sequence":"additional","affiliation":[{"name":"Department of Microbiology, Howard Taylor Ricketts Laboratory, The University of Chicago, Lemont, IL, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zhilei","family":"Chen","sequence":"additional","affiliation":[{"name":"Department of Microbial Pathogenesis and Immunology, Texas A&M College of Medicine, Bryan, TX, USA"},{"name":"Genetics and Genomics Interdisciplinary Program, Texas A&M University, College Station, Texas, USA"},{"name":"Department of Biochemistry and Biophysics, Texas A&M University, College Station, Texas, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2026020510394704000_2026.01.09.698026v2.1","doi-asserted-by":"publisher","DOI":"10.1038\/s41579-018-0147-4"},{"key":"2026020510394704000_2026.01.09.698026v2.2","unstructured":"Siddiqui AH, K.J. Methicillin-Resistant Staphylococcus aureus. (StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing, 2023)."},{"key":"2026020510394704000_2026.01.09.698026v2.3","doi-asserted-by":"crossref","first-page":"97","DOI":"10.1016\/j.cmi.2017.06.017","article-title":"Evaluation of vancomycin MIC creep in methicillin-resistant Staphylococcus aureus infections-a systematic review and meta-analysis","volume":"24","year":"2018","journal-title":"Clinical microbiology and infection : the official publication of the European Society of Clinical Microbiology and Infectious Diseases"},{"key":"2026020510394704000_2026.01.09.698026v2.4","first-page":"3","article-title":"A Canadian perspective on the revised 2020 ASHP-IDSA-PIDS-SIDP guidelines for vancomycin AUC-based therapeutic drug monitoring for serious MRSA infections","volume":"6","year":"2021","journal-title":"J Assoc Med Microbiol Infect Dis Can"},{"key":"2026020510394704000_2026.01.09.698026v2.5","doi-asserted-by":"crossref","first-page":"e0168324","DOI":"10.1128\/spectrum.01683-24","article-title":"Biofilm-mediated antibiotic tolerance in Staphylococcus aureus from spinal cord stimulation device-related infections","volume":"12","year":"2024","journal-title":"Microbiol Spectr"},{"key":"2026020510394704000_2026.01.09.698026v2.6","doi-asserted-by":"crossref","first-page":"6486","DOI":"10.3390\/ijms25126486","article-title":"The Complex Intracellular Lifecycle of Staphylococcus aureus Contributes to Reduced Antibiotic Efficacy and Persistent Bacteremia","volume":"25","year":"2024","journal-title":"International journal of molecular sciences"},{"key":"2026020510394704000_2026.01.09.698026v2.7","doi-asserted-by":"crossref","unstructured":"Streifel, A.C. et al. 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Targeting Alpha Toxin and ClfA with a Multimechanistic Monoclonal-Antibody-Based Approach for Prophylaxis of Serious Staphylococcus aureus Disease. mBio 7 (2016).","DOI":"10.1128\/mBio.00528-16"},{"key":"2026020510394704000_2026.01.09.698026v2.25","doi-asserted-by":"crossref","unstructured":"Tkaczyk, C. et al. Multimechanistic Monoclonal Antibodies (MAbs) Targeting Staphylococcus aureus Alpha-Toxin and Clumping Factor A: Activity and Efficacy Comparisons of a MAb Combination and an Engineered Bispecific Antibody Approach. 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Half-life extension using serum albumin-binding DARPin domains. 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Inhibitory activities of monoclonal antibodies against Staphylococcus aureus clumping factor A. mBio, e0219725 (2025).","DOI":"10.1101\/2025.05.21.655433"},{"key":"2026020510394704000_2026.01.09.698026v2.50","doi-asserted-by":"crossref","unstructured":"Hair, P.S. , Ward, M.D. , Semmes, O.J. , Foster, T.J. & Cunnion, K.M. Staphylococcus aureus clumping factor A binds to complement regulator factor I and increases factor I cleavage of C3b. J Infect Dis 1G8, 125\u2013133 (2008).","DOI":"10.1086\/588825"},{"key":"2026020510394704000_2026.01.09.698026v2.51","doi-asserted-by":"publisher","DOI":"10.1128\/IAI.01065-09"},{"key":"2026020510394704000_2026.01.09.698026v2.52","doi-asserted-by":"publisher","DOI":"10.1038\/nprot.2007.132"},{"key":"2026020510394704000_2026.01.09.698026v2.53","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M110.214981"},{"key":"2026020510394704000_2026.01.09.698026v2.54","doi-asserted-by":"crossref","unstructured":"Schlothauer, T. et al. 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