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Svante Eriksson (Institutionen för fysikalisk kemi) ; Bengt Nordén (Institutionen för fysikalisk kemi) ; Katsumi Morimatsu ; T. Horii ; Masayuki Takahashi
Journal of Biological Chemistry (0021-9258). Vol. 268 (1993), 3, p. 1811-1816.
[Artikel, refereegranskad vetenskaplig]

The tyrosine fluorescence of the RecA protein is quenched by about 15% upon binding of the cofactor analog adenosine 5'-O-(3-thiotriphosphate) (ATPgammaS). This quenching is not observed with a modified RecA in which the tyrosine residue at position 264 (Tyr-264) is replaced for alanine by site-directed mutagenesis, a modification which also results in a decrease of binding affinity of cofactor. This indicates that Tyr-264 is responsible for the fluorescence change and that the residue is close to or within the cofactor binding site. Upon DNA binding, a change of tyrosine fluorescence is observed both with the modified protein and with wild type RecA, indicating that DNA binding affects the environment of other tyrosine residues than Tyr-264. However, the change is significantly smaller in the modified protein, suggesting that both Tyr-264 as well as other residue(s) may be affected by the DNA binding. Changed fluorescence properties of the remaining tyrosine residues as a result of a slightly different DNA binding mode of the modified protein are also possible. Tyr-264 may be an important residue for the allosteric effect induced by the cofactor for the binding of DNA to RecA. In the recent crystal structure of RecA-ADP published by Story and Steitz (Story, R. M., and Steitz, T. A. (1992) Nature 355, 374-376), ADP is stacked with Tyr-103 and does not interact with Tyr-264. The fact that we observe no interaction of ATPgammaS with Tyr-103 (as evidenced from absence of fluorescence change) but instead with Tyr-264 may suggest an important conformational difference between the RecA complexes with, respectively, ADP and ATP.

Nyckelord: single-stranded-dna, escherichia-coli, linear dichroism, general, recombination, electron-microscopy, nucleotide cofactor, lambda-repressor, atp hydrolysis, protein, complexes

Denna post skapades 2011-08-17.
CPL Pubid: 144385


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