Lin YC, et al. (2007) Genetic analysis reveals essential and non-essential amino acids within the telomeric DNA-binding interface of Cdc13p. Biochem J 403(2):289-95
Abstract: Cdc13p is a specific single-stranded telomeric DNA binding protein of Saccharomyces cerevisiae. It is involved in protecting telomeres and regulating telomere length. The telomere binding domain of Cdc13p is located within residues 497-693 and its structure has been resolved by NMR spectroscopy. A series of aromatic, hydrophobic, and basic residues located at the DNA-binding surface of Cdc13p are involved for binding to telomeres. Here we applied a genetic approach to analyze the involvements of these residues in telomere binding. A series of mutants within the telomere binding domain of Cdc13p were identified that failed to complement cdc13 mutants in vivo. Among the amino acids residues that were isolated, the Y522, R635, and I633 residues were shown to locate at the DNA binding surface. We further demonstrated that Y522C and R635A mutants failed to bind telomeric DNA in vitro, indicating that these residues are indeed required for telomere binding. We did not, however, isolate other mutant residues located at the DNA binding surface of Cdc13p beyond these three residues. Instead, a mutant on K568 was isolated that did not affect the essential function of Cdc13p. The K568 is also located on the DNA binding surface of Cdc13p. Thus, these results suggested that other DNA binding residues are not essential for telomere binding. In sum, we have established a genetic test that enabled the identification of telomere-binding residues of Cdc13p in vivo. This type of analysis provides information on which residues are indeed contributes to telomere binding in vivo.
|Status: Published||Type: Journal Article||PubMed ID: 17166094|
Topics addressed in this paper
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|Protein Sequence Features|
|Protein-Nucleic Acid Interactions|