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Roles of Asp54 and Asp213 in Ca2+ utilization by soluble human CD39/ecto-nucleotidase.

Authors
  • Drosopoulos, Joan H F
Type
Published Article
Journal
Archives of biochemistry and biophysics
Publication Date
Oct 01, 2002
Volume
406
Issue
1
Pages
85–95
Identifiers
PMID: 12234494
Source
Medline
License
Unknown

Abstract

Soluble human CD39 (solCD39) rapidly metabolizes nucleotides, especially ADP released from activated platelets, thereby inhibiting further platelet activation and recruitment. Using alanine substitution mutagenesis, we established a functional role for aspartates D54 and D213 in solCD39. Kinetic analyses of D54A and D213A indicated decreased K(m)s of the mutants, compared to wild type, for the cofactor calcium and for the substrates ADP and ATP. These decreases in calcium and nucleotide affinity of the mutants were accompanied by increases in their rate of catalysis. The decreased affinity of the mutants for calcium was responsible for their diminished ability to reverse platelet aggregation in plasma anticoagulated with citrate, a known calcium chelator. Their ADPase activity in the presence of citrated plasma was also decreased, although this could be overcome with excess calcium. Thus, aspartates 54 and 213 are involved in calcium utilization and potentially involved in cation coordination with substrate in the catalytic pocket of solCD39.

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