4GIJ

Crystal Structure of Pseudouridine Monophosphate Glycosidase Complexed with Sulfate

Summary for 4GIJ

• Entry DOI: 10.2210/pdb4gij/pdb
• Related: 4GIK 4GIL 4GIM
• Descriptor: Pseudouridine-5'-phosphate glycosidase, SULFATE ION, MANGANESE (II) ION, ... (4 entities in total)
• Functional Keywords: alpha-beta-alpha sandwich fold, hydrolase
• Biological source: Escherichia coli
• Total number of polymer chains: 3
• Total formula weight: 107323.96

Authors

Huang, S.,Mahanta, N.,Begley, T.P.,Ealick, S.E. (deposition date: 2012-08-08, release date: 2012-10-31, Last modification date: 2023-09-13)

Primary citation

Huang, S.,Mahanta, N.,Begley, T.P.,Ealick, S.E.
Pseudouridine monophosphate glycosidase: a new glycosidase mechanism.
Biochemistry, 51:9245-9255, 2012

PubMed Abstract: Pseudouridine (Ψ), the most abundant modification in RNA, is synthesized in situ using Ψ synthase. Recently, a pathway for the degradation of Ψ was described [Preumont, A., Snoussi, K., Stroobant, V., Collet, J. F., and Van Schaftingen, E. (2008) J. Biol. Chem. 283, 25238-25246]. In this pathway, Ψ is first converted to Ψ 5'-monophosphate (ΨMP) by Ψ kinase and then ΨMP is degraded by ΨMP glycosidase to uracil and ribose 5-phosphate. ΨMP glycosidase is the first example of a mechanistically characterized enzyme that cleaves a C-C glycosidic bond. Here we report X-ray crystal structures of Escherichia coli ΨMP glycosidase and a complex of the K166A mutant with ΨMP. We also report the structures of a ring-opened ribose 5-phosphate adduct and a ring-opened ribose ΨMP adduct. These structures provide four snapshots along the reaction coordinate. The structural studies suggested that the reaction utilizes a Lys166 adduct during catalysis. Biochemical and mass spectrometry data further confirmed the existence of a lysine adduct. We used site-directed mutagenesis combined with kinetic analysis to identify roles for specific active site residues. Together, these data suggest that ΨMP glycosidase catalyzes the cleavage of the C-C glycosidic bond through a novel ribose ring-opening mechanism.

PubMed: 23066817
DOI: 10.1021/bi3006829

Experimental method

X-RAY DIFFRACTION (1.941 Å)