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Closed Access item Overproduction, purification, and characterization of DNA-binding protein P19 of bacteriophage PRD1

Authors:Pakula, Tiina M.
Caldentey, Javier
Gutiérrez Armenta, Crisanto
Olkkonen, Vesa M.
Bamford, Dennis H.
Salas, Margarita
Keywords:Protection of single-stranded DNA against nuclease P1, Phage λ pL promoter, Protein purification, Gel-shift assay
Issue Date:15-Apr-1993
Publisher:Elsevier
Citation:Gene 126(1): 99-104 (1993)
Abstract:The early protein, P 19, of bacteriophage PRD1 was purified after overexpression of the cloned gene, XIX, in Escherichia coli DH5α cells. The purified protein binds as multimers to single-stranded DNA (ssDNA), and with a lower affinity to double-stranded DNA (dsDNA), without sequence-specificity. Two distinct P19-ssDNA complexes were discovered in gel- mobility-shift assays at different protein:DNA ratios. P 19 was capable of fully protecting ssDNA against nuclease P1. Electron microscopy of protein P19-ssDNA complexes showed DNA molecules which were extensively coated with protein and whose contour length was clearly reduced by P 19 binding. The results suggest that P 19 binds to ssDNA with moderate cooperativity and are consistent with the DNA being wrapped around the P 19 multimers.
Publisher version (URL):http://dx.doi.org/10.1016/0378-1119(93)90595-T
URI:http://hdl.handle.net/10261/39360
ISSN:0378-1119
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