{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T02:53:22Z","timestamp":1760151202819,"version":"build-2065373602"},"reference-count":32,"publisher":"MDPI AG","issue":"2","license":[{"start":{"date-parts":[[2022,2,10]],"date-time":"2022-02-10T00:00:00Z","timestamp":1644451200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100000060","name":"National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["R01AI150480"],"award-info":[{"award-number":["R01AI150480"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Service de Sant\u00e9 des Arm\u00e9es and the D\u00e9l\u00e9gation G\u00e9n\u00e9rale pour l\u2032Armement","award":["No number"],"award-info":[{"award-number":["No number"]}]},{"name":"French National Research Agency in the framework of the &quot;Investissements d\u2019avenir\u201d program","award":["ANR-15-IDEX-02"],"award-info":[{"award-number":["ANR-15-IDEX-02"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Viruses"],"abstract":"<jats:p>A modified SELEX (Systematic Evolution of Ligands by Exponential Enrichment) pr,otocol (referred to as PT SELEX) was used to select primer\u2013template (P\/T) sequences that bound to the vaccinia virus polymerase catalytic subunit (E9) with enhanced affinity. A single selected P\/T sequence (referred to as E9-R5-12) bound in physiological salt conditions with an apparent equilibrium dissociation constant (KD,app) of 93 \u00b1 7 nM. The dissociation rate constant (koff) and binding half-life (t1\/2) for E9-R5-12 were 0.083 \u00b1 0.019 min\u22121 and 8.6 \u00b1 2.0 min, respectively. The values indicated a several-fold greater binding ability compared to controls, which bound too weakly to be accurately measured under the conditions employed. Loop-back DNA constructs with 3\u2032-recessed termini derived from E9-R5-12 also showed enhanced binding when the hybrid region was 21 nucleotides or more. Although the sequence of E9-R5-12 matched perfectly over a 12-base-pair segment in the coding region of the virus B20 protein, there was no clear indication that this sequence plays any role in vaccinia virus biology, or a clear reason why it promotes stronger binding to E9. In addition to E9, five other polymerases (HIV-1, Moloney murine leukemia virus, and avian myeloblastosis virus reverse transcriptases (RTs), and Taq and Klenow DNA polymerases) have demonstrated strong sequence binding preferences for P\/Ts and, in those cases, there was biological or potential evolutionary relevance. For the HIV-1 RT, sequence preferences were used to aid crystallization and study viral inhibitors. The results suggest that several other DNA polymerases may have P\/T sequence preferences that could potentially be exploited in various protocols.<\/jats:p>","DOI":"10.3390\/v14020369","type":"journal-article","created":{"date-parts":[[2022,2,11]],"date-time":"2022-02-11T02:40:17Z","timestamp":1644547217000},"page":"369","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["Selection of Primer\u2013Template Sequences That Bind with Enhanced Affinity to Vaccinia Virus E9 DNA Polymerase"],"prefix":"10.3390","volume":"14","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-8710-7622","authenticated-orcid":false,"given":"Jeffrey J.","family":"DeStefano","sequence":"first","affiliation":[{"name":"Department of Cell Biology and Molecular Genetics, Bioscience Research Building, University of Maryland, College Park, MD 20742, USA"},{"name":"Maryland Pathogen Research Institute (MPRI), University of Maryland, College Park, MD 20742, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Fr\u00e9d\u00e9ric","family":"Iseni","sequence":"additional","affiliation":[{"name":"Unit\u00e9 de Virologie, Institut de Recherche Biom\u00e9dicale des Arm\u00e9es, BP73, 91223 Br\u00e9tigny-sur-Orge, France"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Nicolas","family":"Tarbouriech","sequence":"additional","affiliation":[{"name":"Institut de Biologie Structurale, Universit\u00e9 Grenoble Alpes, Centre National de la Recherche Scientifique, Commissariat \u00e0 l\u2019Energie Atomique et aux Energies Alternatives, IBS, 38000 Grenoble, France"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2022,2,10]]},"reference":[{"key":"ref_1","first-page":"187","article-title":"Adenovirus DNA Replication: Protein Priming, Jumping Back and the Role of the DNA Binding Protein DBP","volume":"272","author":"Brenkman","year":"2003","journal-title":"Curr. 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