3AO9

Crystal structure of the C-terminal domain of sequence-specific ribonuclease

Summary for 3AO9

• Entry DOI: 10.2210/pdb3ao9/pdb
• Related: 2DJH
• Descriptor: Colicin-E5, CADMIUM ION (3 entities in total)
• Functional Keywords: ribonuclease, hydrolase
• Biological source: Escherichia coli
• Total number of polymer chains: 2
• Total formula weight: 26158.06

Authors

Inoue, S.,Fushinobu, S.,Ogawa, T.,Hidaka, M.,Masaki, H.,Yajima, S. (deposition date: 2010-09-22, release date: 2011-09-28, Last modification date: 2023-11-01)

Primary citation

Inoue-Ito, S.,Yajima, S.,Fushinobu, S.,Nakamura, S.,Ogawa, T.,Hidaka, M.,Masaki, H.
Identification of the catalytic residues of sequence-specific and histidine-free ribonuclease colicin E5
J.Biochem., 152:365-372, 2012

PubMed Abstract: Colicin E5 cleaves tRNAs for Tyr, His, Asn and Asp in their anticodons to abolish protein synthesis in Escherichia coli. We previously showed how its C-terminal RNase domain, E5-CRD, recognizes the anticodon bases but the catalytic mechanism remained to be elucidated. Although the reaction products with 5'-OH and 2',3'-cyclic phosphate ends suggested a similar mechanism to those of RNases A and T1, E5-CRD does not have the His residues necessary as a catalyst in usual RNases. To identify residues important for the catalytic reaction, mutants as to all residues within 5 Å from the central phosphorus of the scissile phosphodiester bond were prepared. Evaluation of the killing activities of the mutant colicins and the RNase activities of the mutant E5-CRDs suggested direct involvement of Arg33, Lys25, Gln29 and Lys60 in the reaction. Particularly, Arg33 plays a critical role and Ile94 provides a structural support of Arg33. Crystal structure of the complex of E5-CRD(R33Q)/dGpdUp showed structural and binding functional integrity of this mutant protein, suggesting involvement of Arg33 in the catalytic reaction. The structure of the free E5-CRD, we also determined, showed great flexibility of a flap region, which facilitates the access of Lys60 to the substrate in an induced-fit manner.

PubMed: 22815490
DOI: 10.1093/jb/mvs077

Experimental method

X-RAY DIFFRACTION (2.1 Å)