The nucleotide pyrophosphatases/phosphodiesterases NPP1 and NPP2/autotaxin are structurally related eukaryotic ecto-enzymes, but display a very different substrate specificity. NPP1 releases nucleoside 5′-monophosphates from various nucleotides, whereas NPP2 mainly functions as a lysophospholipase D. We have used a domain-swapping approach to map substrate-specifying determinants of NPP1 and NPP2. The catalytic domain of NPP1 fused to the N- and C-terminal domains of NPP2 was hyperactive as a nucleotide phosphodiesterase, but did not show any lysophospholipase D activity. In contrast, chimaeras of the catalytic domain of NPP2 and the N- and/or C-terminal domains of NPP1 were completely inactive. These data indicate that the catalytic domain as well as both extremities of NPP2 contain lysophospholipid-specifying sequences. Within the catalytic domain of NPP1 and NPP2, we have mapped residues close to the catalytic site that determine the activities towards nucleotides and lysophospholipids. We also show that the conserved Gly/Phe-Xaa-Gly-Xaa-Xaa-Gly (G/FXGXXG) motif near the catalytic site is required for metal binding, but is not involved in substrate-specification. Our data suggest that the distinct activities of NPP1 and NPP2 stem from multiple differences throughout the polypeptide chain.
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Research Article|
June 22 2004
Substrate-specifying determinants of the nucleotide pyrophosphatases/phosphodiesterases NPP1 and NPP2
Anisoara CIMPEAN;
Anisoara CIMPEAN
1Afdeling Biochemie, Faculteit Geneeskunde, Katholieke Universiteit Leuven, Herestraat 49, B-3000 Leuven, Belgium
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Cristiana STEFAN;
Cristiana STEFAN
1Afdeling Biochemie, Faculteit Geneeskunde, Katholieke Universiteit Leuven, Herestraat 49, B-3000 Leuven, Belgium
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Rik GIJSBERS;
Rik GIJSBERS
1Afdeling Biochemie, Faculteit Geneeskunde, Katholieke Universiteit Leuven, Herestraat 49, B-3000 Leuven, Belgium
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Willy STALMANS;
Willy STALMANS
1Afdeling Biochemie, Faculteit Geneeskunde, Katholieke Universiteit Leuven, Herestraat 49, B-3000 Leuven, Belgium
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Mathieu BOLLEN
Mathieu BOLLEN
1
1Afdeling Biochemie, Faculteit Geneeskunde, Katholieke Universiteit Leuven, Herestraat 49, B-3000 Leuven, Belgium
1To whom correspondence should be addressed (e-mail Mathieu.Bollen@med.kuleuven.ac.be).
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Publisher: Portland Press Ltd
Received:
March 22 2004
Revision Received:
April 14 2004
Accepted:
April 20 2004
Accepted Manuscript online:
April 20 2004
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London
2004
Biochem J (2004) 381 (1): 71–77.
Article history
Received:
March 22 2004
Revision Received:
April 14 2004
Accepted:
April 20 2004
Accepted Manuscript online:
April 20 2004
Connected Content
A correction has been published:
Correction: Substrate-specifying determinants of the nucleotide pyrophosphatases/phosphodiesterases NPP1 and NPP2
Citation
Anisoara CIMPEAN, Cristiana STEFAN, Rik GIJSBERS, Willy STALMANS, Mathieu BOLLEN; Substrate-specifying determinants of the nucleotide pyrophosphatases/phosphodiesterases NPP1 and NPP2. Biochem J 1 July 2004; 381 (1): 71–77. doi: https://doi.org/10.1042/BJ20040465
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