Enzymatic activation of autotaxin by divalent cations without EF-hand loop region involvement.

Abstract

Autotaxin (ATX) is a recently described member of the nucleotide pyrophosphatase/phosphodiesterase (NPP) family of proteins with potent tumor cell motility-stimulating activity. Like other NPPs, ATX is a glycoprotein with peptide sequences homologous to the catalytic site of bovine intestinal alkaline phosphodiesterase (PDE) and the loop region of an EF-hand motif. The PDE active site of ATX has been associated with the motility-stimulating activity of ATX. In this study, we examined the roles of the EF-hand loop region and of divalent cations on the enzymatic activities of ATX. Ca(2+) or Mg(2+) was each demonstrated to increase the PDE activity of ATX in a concentration-dependent manner, whereas incubation of ATX with chelating agents abolished this activity, indicating a requirement for divalent cations. Non-linear regression analysis of enzyme kinetic data indicated that addition of these divalent cations increases reaction velocity predominantly through an effect on V(max.) Three mutant proteins, Ala(740)-, Ala(742)-, and Ala(751)-ATX, in the EF-hand loop region of ATX had enzymatic activity comparable to that of the wild-type protein. A deletion mutation of the entire loop region resulted in slightly reduced PDE activity but normal motility-stimulating activity. However, the PDE activity of this same deletion mutant remained sensitive to augmentation by cations, strongly implying that cations exert their effect by interactions outside of the EF-hand loop region.

• Authors: J Lee, I D Jung, S W Nam, T Clair, E M Jeong, S Y Hong, J W Han, H W Lee, M L Stracke, H Y Lee
• Journal: Biochemical pharmacology (Biochem Pharmacol) Vol. 62 Issue 2 Pg. 219-24 (Jul 15 2001) ISSN: 0006-2952 [Print] England
• PMID: 11389881 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Cations, Divalent
• Chelating Agents
• Glycoproteins
• Multienzyme Complexes
• Phosphoric Diester Hydrolases
• Phosphodiesterase I
• alkylglycerophosphoethanolamine phosphodiesterase
• Pyrophosphatases
• nucleotide pyrophosphatase - phosphodiesterase I
• Glucose-6-Phosphate Isomerase

Topics

• Amino Acid Sequence
• Animals
• Biotransformation (drug effects)
• COS Cells
• Cations, Divalent (pharmacology)
• Chelating Agents (pharmacology)
• Glucose-6-Phosphate Isomerase (metabolism)
• Glycoproteins (metabolism)
• Molecular Sequence Data
• Multienzyme Complexes
• Phosphodiesterase I
• Phosphoric Diester Hydrolases (metabolism)
• Protein Structure, Tertiary
• Pyrophosphatases (metabolism)
• Sequence Homology, Amino Acid