Cloning, expression, and functional characterization of a Ca(2+)-dependent endoplasmic reticulum nucleoside diphosphatase

J Biol Chem. 2002 Oct 4;277(40):36978-86. doi: 10.1074/jbc.M201656200. Epub 2002 Aug 6.


We have isolated and characterized the cDNA encoding a Ca(2+)-dependent nucleoside diphosphatase (EC ) related to two secreted ATP- and ADP-hydrolyzing apyrases of the bloodsucking insects, Cimex lectularius and Phlebotomus papatasi. The rat brain-derived cDNA has an open reading frame of 1209 bp encoding a protein of 403 amino acids and a calculated molecular mass of 45.7 kDa. The mRNA was expressed in all tissues investigated, revealing two major transcripts with varying preponderance. The immunohistochemical analysis of the Myc-His-tagged enzyme expressed in Chinese hamster ovary cells revealed its association with the endoplasmic reticulum and also with pre-Golgi intermediates. Ca(2+)-dependent nucleoside diphosphatase is a membrane protein with its catalytic site facing the organelle lumen. It hydrolyzes nucleoside 5'-diphosphates in the order UDP >GDP = IDP >>>CDP but not ADP. Nucleoside 5'-triphosphates were hydrolyzed to a minor extent, and no hydrolysis of nucleoside 5'-monophosphates was observed. The enzyme was strongly activated by Ca(2+), insensitive to Mg(2+), and had a K(m) for UDP of 216 microm. Ca(2+)-dependent nucleoside diphosphatase may support glycosylation reactions related to quality control in the endoplasmic reticulum.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acid Anhydride Hydrolases / chemistry
  • Acid Anhydride Hydrolases / genetics*
  • Acid Anhydride Hydrolases / metabolism
  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Brain / enzymology*
  • CHO Cells
  • Calcium / physiology*
  • Cells, Cultured
  • Cloning, Molecular
  • Cricetinae
  • Cytosol / enzymology
  • DNA Primers
  • Endoplasmic Reticulum / physiology*
  • Escherichia coli / enzymology
  • Escherichia coli / genetics
  • Gene Library
  • Humans
  • Molecular Sequence Data
  • Polymerase Chain Reaction
  • Protein Conformation
  • Rats
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / metabolism
  • Reverse Transcriptase Polymerase Chain Reaction
  • Transfection


  • DNA Primers
  • Recombinant Proteins
  • Acid Anhydride Hydrolases
  • nucleoside-diphosphatase
  • Calcium

Associated data

  • GENBANK/AJ312207
  • GENBANK/AJ312208