2C06

NMR-based model of the complex of the toxin Kid and a 5-nucleotide substrate RNA fragment (AUACA)

Summary for 2C06

• Entry DOI: 10.2210/pdb2c06/pdb
• Related: 1M1F
• NMR Information: BMRB: 6925
• Descriptor: KID TOXIN PROTEIN, 5'-R(*AP*UP*AP*CP*AP)-3' (2 entities in total)
• Functional Keywords: docking, dna replication, mazf, plasmid maintenance, post segregational killing, protein-rna complex, ribonuclease, rna cleavage, rnase, toxin-antitoxin, toxin
• Biological source: ESCHERICHIA COLI
• Total number of polymer chains: 3
• Total formula weight: 25345.53

Authors

Kamphuis, M.B.,Bonvin, A.M.J.J.,Monti, M.C.,Lemonnier, M.,Munoz-Gomez, A.,Van Den Heuvel, R.H.H.,Diaz-Orejas, R.,Boelens, R. (deposition date: 2005-08-25, release date: 2006-02-08, Last modification date: 2024-06-19)

Primary citation

Kamphuis, M.B.,Bonvin, A.M.J.J.,Monti, M.C.,Lemonnier, M.,Munoz-Gomez, A.,Van Den Heuvel, R.H.H.,Diaz-Orejas, R.,Boelens, R.
Model for RNA Binding and the Catalytic Site of the Rnase Kid of the Bacterial Pard Toxin-Antitoxin System.
J.Mol.Biol., 357:115-, 2006

PubMed Abstract: The toxin Kid and antitoxin Kis are encoded by the parD operon of Escherichia coli plasmid R1. Kid and its chromosomal homologues MazF and ChpBK have been shown to inhibit protein synthesis in cell extracts and to act as ribosome-independent endoribonucleases in vitro. Kid cleaves RNA preferentially at the 5' side of the A residue in the nucleotide sequence 5'-UA(A/C)-3' of single-stranded regions. Here, we show that RNA cleavage by Kid yields two fragments with a 2':3'-cyclic phosphate group and a free 5'-OH group, respectively. The cleavage mechanism is similar to that of RNases A and T1, involving the uracil 2'-OH group. Via NMR titration studies with an uncleavable RNA mimic, we demonstrate that residues of both monomers of the Kid dimer together form a concatenated RNA-binding surface. Docking calculations based on the NMR chemical shifts, the cleavage mechanism and previously reported mutagenesis data provide a detailed picture of the position of the AUACA fragment within the binding pocket. We propose that residues D75, R73 and H17 form the active site of the Kid toxin, where D75 and R73 are the catalytic base and acid, respectively. The RNA sequence specificity is defined by residues T46, S47, A55, F57, T69, V71 and R73. Our data show the importance of these residues for Kid function, and the implications of our results for related toxins, such as MazF, CcdB and RelE, are discussed.

PubMed: 16413033
DOI: 10.1016/J.JMB.2005.12.033

Experimental method

SOLUTION NMR