Repressor-operator interaction in the lac operon: III. Nuclear magnetic resonance observations with altered amino-terminal DNA binding domains |
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Authors: | Kim Arndt Harry Nick Frank Boschelli Ponzy Lu John Sadler |
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Institution: | Department of Chemistry, University of Pennsylvania Philadelphia, PA 19104, U.S.A.;Department of Biophysics and Genetics University of Colorado Health Science Center Denver, CO 80262, U.S.A. |
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Abstract: | We have examined the N-terminal 56 amino acid fragment, the domain that can bind DNA independently, from 3-fluorotyrosine-substituted Escherichia coli lac repressor by 19F-nuclear magnetic resonance. The fragments or “headpieces” from four altered repressers missing each of the tyrosines in turn were examined in parallel. When the wild-type N-terminal fragment is titrated with a 36 base-pair lac operator DNA sequence, the 19F resonances undergo changes in their chemical shifts that are different from those changes when the N-terminal fragment is titrated with non-specific DNA fragments. By looking at these operator-induced changes as well as pH-dependent effects with all four altered N-terminal fragments, we show systematic correlations with the genetic data. The data lead us to conclude that upon operator DNA binding: (1) tyrosine 7 is displaced to a less polar environment and the higher than normal pK value of the phenolic OH group is decreased; (2) tyrosine 12 does not change much in either its mobility or environment; and (3) tyrosine 17 is involved, as suggested by the genetic data, when the headpiece forms a complex with operator DNA. |
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Keywords: | n m r nuclear magnetic resonance |
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