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EMBO J. 2000 Nov 15;19(22):6207-17.

Structure and mechanism of activity of the cyclic phosphodiesterase of Appr>p, a product of the tRNA splicing reaction.

Author information

1
Protein Structure Section, Macromolecular Crystallography Laboratory, Program in Structural Biology, NCI-Frederick, Frederick, MD 21702, USA. hofmanna@ncifcrf.gov

Abstract

The crystal structure of the cyclic phosphodiesterase (CPDase) from Arabidopsis thaliana, an enzyme involved in the tRNA splicing pathway, was determined at 2.5 A resolution. CPDase hydrolyzes ADP-ribose 1",2"-cyclic phosphate (Appr>p), a product of the tRNA splicing reaction, to the monoester ADP-ribose 1"-phosphate (Appr-1"p). The 181 amino acid protein shows a novel, bilobal arrangement of two alphabeta modules. Each lobe consists of two alpha-helices on the outer side of the molecule, framing a three- or four-stranded antiparallel beta-sheet in the core of the protein. The active site is formed at the interface of the two beta-sheets in a water-filled cavity involving residues from two H-X-T/S-X motifs. This previously noticed motif participates in coordination of a sulfate ion. A solvent-exposed surface loop (residues 100-115) is very likely to play a flap-like role, opening and closing the active site. Based on the crystal structure and on recent mutagenesis studies of a homologous CPDase from Saccharomyces cerevisiae, we propose an enzymatic mechanism that employs the nucleophilic attack of a water molecule activated by one of the active site histidines.

PMID:
11080166
PMCID:
PMC305825
DOI:
10.1093/emboj/19.22.6207
[Indexed for MEDLINE]
Free PMC Article

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