In eukaryotes, nucleotide excision repair of ultraviolet light-damaged DNA is a highly intricate process that requires a large number of evolutionarily conserved protein factors. Genetic studies in the yeast Saccharomyces cerevisiae have indicated a specific role of the RAD7 and RAD16 genes in the repair of transcriptionally inactive DNA. Here we show that the RAD7- and RAD16-encoded products exist as a complex of 1:1 stoichiometry, exhibiting an apparent dissociation constant (Kd) of <4 x 10(-10) M. The Rad7-Rad16 complex has been purified to near homogeneity in this study and is shown to bind, in an ATP-dependent manner and with high specificity, to DNA damaged by ultraviolet light. Importantly, inclusion of the Rad7-Rad16 complex in the in vitro nucleotide excision repair system that consists entirely of purified components results in a marked stimulation of damage specific incision. Thus, Rad7-Rad16 complex is the ATP-dependent DNA damage sensor that specifically functions with the ensemble of nucleotide excision repair factor (NEF) 1, NEF2, NEF3, and replication protein A in the repair of transcriptionally inactive DNA. We name this novel complex of Rad7 and Rad16 proteins NEF4.

• PMID: 9268290
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Journal Article | Research Support, U.S. Gov't, Non-P.H.S. | Research Support, U.S. Gov't, P.H.S.

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Guzder SN, Sung P, Prakash L, Prakash S

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