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Identification of the substrate binding site in the N‐terminal TBP‐like domain of RNase H3
Author(s) -
Miyashita Seiko,
Tadokoro Takashi,
Angkawidjaja Clement,
You Dong-Ju,
Koga Yuichi,
Takano Kazufumi,
Kanaya Shigenori
Publication year - 2011
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/j.febslet.2011.05.064
Subject(s) - rnase p , substrate (aquarium) , rnase h , binding site , binding domain , ribonuclease , chemistry , dna , biochemistry , s tag , biophysics , biology , stereochemistry , rna , ecology , gene
Ribonuclease H3 from Bacillus stearothermophilus (Bst‐RNase H3) has the N‐terminal TBP‐like substrate‐binding domain. To identify the substrate binding site in this domain, the mutant proteins of the intact protein and isolated N‐domain, in which six of the seventeen residues corresponding to those involved in DNA binding of TBP are individually mutated to Ala, were constructed. All of them exhibited decreased enzymatic activities and/or substrate‐binding affinities when compared to those of the parent proteins, suggesting that the N‐terminal domain of RNase H3 uses the flat surface of the β‐sheet for substrate binding as TBP to bind DNA. This domain may greatly change conformation upon substrate binding.