A boundary sedimentation velocity method for determining nonspecific nucleic acid-protein interaction binding parameters

✍ Scribed by David E. Jensen; Peter H. von Hippel

• Publisher: Elsevier Science
• Year: 1977
• Tongue: English
• Weight: 961 KB
• Volume: 80
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(77)90645-5

✦ Synopsis

An absolute method for measuring affinity constants and binding site sizes for the non-sequence-specific binding of protein to single-or double-stranded polynucleotide chains is described, based on the difference in the rate of sedimentation of free protein and the protein-nucleic acid complexes. In the form outlined, the technique can measure binding constants ranging from 103 to 106 M-'; methods for extending the range of measurable binding constants are also described. Data obtained for the noncooperative binding of ribonuclease to DNA are presented as fully worked-out examples of the method, and the extension of this approach to other noncooperative and cooperative protein-nucleic acid binding systems is discussed.