Biomedical subjects

S S York

Publications and source records attributed to S S York.

3 recordsLinked to original sources

PubMed1979

lac repressor changes conformation upon binding to poly[dA-T)].

N-(Iodoacetylaminoethyl)-1-naphthylamine-5-sulfonate reacts with Escherichia coli lac repressor to selectively label cysteine-140 with the fluorescent N-(acetylaminoethyl)-1-naphthylamine-5-sulfonate group. The fluorescence intensity of this label decreases by 20% when labeled repressor associates with poly[d(A-T)]. Fifteen base pairs of poly[d(A-T)] per repressor tetramer are required to complete this decrease. Stopped-flow experiments have shown that the repressor undergoes at least two conformational changes as it binds to poly[d(A-T)], with half-lives of 5.0 +/- 1.2 msec and 3.5 +/- 1.0 sex. Quite likely, these conformational changes serve to strengthen the interaction of repressor with DNA.

Cysteine↗

PubMed1978

Binding of recrystallized and chromatographically purified 8-anilino-1-naphthalenesulfonate to Escherichia coli lac repressor.

8-Anilion-1-naphthalenesulfonate (Ans), recrystallized from water as the magnesium salt, contains a fluorescent impurity representing 0.3% of the absorbance at 351 nm. This impurity can be removed by Sephadex LH-20 chromatography. The chromatographic and spectral properties of this impurity suggest that it is bis(Ans), a dimer of Ans. This bis(Ans) impurity makes a significant contribution to the fluorescence increment observed when lac repressor is added to recrystallized Ans. This occurs because bis(Ans) binds much more tightly to this protein than does Ans. The dissociation constant divided by the number of binding sites per subunit is 3.1 X 10(-6) M for bis(Ans); the corresponding value for Ans is greater than 1 X 10(-4) M. Because of their differing absorption spectra, the impact of this bis(Ans) impurity is especially large with excitation wavelengths above 400 nm. Users of recrystallized Ans should consider the potential consequences of this impurity whenever working with a protein to which Ans binds weakly.

Anilino Naphthalenesulfonates↗

PubMed1978

Association of Escherichia coli lac repressor with poly[d(A-T)] monitored with 8-anilino-1-napthalenesulfonate.

The association of lac repressor with poly[d(A-T)] was monitored with the fluorescent prob 8-anilino-1-naphthalenesulfonate (Ans). Excess poly[d(A-T)] decreased the emission intensity of the repressor--Ans complex by 30%. Fluorescence titrations indicated that 33 +/- 4 base pairs were required to bind all of the repressor. Sedimentation studies indicated, however, that all of the repressor sedimented as a protein--DNA complex with as few as 10 to 15 base pairs per tetramer, even in the presence of Ans. These data are interpreted with two models: one where repressors bind to both sides of the DNA (Butler, A. P., et al. (1977) Biochemistry 16, 4757: Zingsheim, H.P., et al. (1977) J. Mol. Biol. 115, 565), the other where a double layer of repressors bind to a single side of the DNA. Removal of the amino-terminal regions from the repressor decreased the fluorescence from bound Ans by 77%. The amino-terminal fragments alone did not enhance Ans fluorescence.

Anilino Naphthalenesulfonates↗