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ADP-ribosyl cyclase; crystal structures reveal a covalent intermediate
Michael L Love1, Doletha M E Szebenyi, Irina A Kriksunov
1Molecular Biology and Genetics, Cornell University, Ithaca, NY 14853 USA.
Structure (London, England : 1993)
|March 16, 2004
Summary
Aplysia cyclase
Area of Science:
- Biochemistry
- Structural Biology
- Enzymology
Background:
- ADP-ribosyl cyclase enzymes are crucial for synthesizing cyclic ADP-ribose (cADPR), a vital second messenger in calcium signaling.
- Understanding the structural basis of cyclase activity is key to elucidating cellular calcium regulation.
Purpose of the Study:
- To determine the high-resolution crystal structures of Aplysia ADP-ribosyl cyclase in complex with various substrates and inhibitors.
- To elucidate the catalytic mechanism and identify key active site features involved in NAD+ cyclization.
Main Methods:
- X-ray crystallography was employed to resolve the structures of Aplysia cyclase at resolutions up to 2.0 Å.
- Covalent adducts with ribose-5-phosphate and pyridylcarbinol, as well as nicotinamide-bound states, were analyzed.
- Structures of both native and Glu179Ala mutant cyclase were determined.
Main Results:
- The structure revealed covalent binding of ribose-5-phosphate and pyridylcarbinol at the active site.
- A second nicotinamide binding site was identified in the nicotinamide-bound cyclase structure.
- The Glu179Ala mutation provided insights into the enzyme's catalytic role.
Conclusions:
- A proposed mechanism suggests a second nicotinamide site aids cyclization by clearing the nicotinamide byproduct.
- The findings support a ribosylation mechanism involving a covalently bound intermediate.
- These structural insights advance the understanding of cADPR synthesis and calcium signaling pathways.