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

Lee, Jangsoon; Jung, In Duk; Nam, Suk Woo; Clair, Tim; Jeong, Eui Myung; Hong, Sung Yeol; Han, Jeung Whan; Lee, Hyang Woo; Stracke, Mary L.; Lee, Hoi Young

doi:10.1016/s0006-2952(01)00658-x

Cited by 25 publications

(8 citation statements)

References 21 publications

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“…The NUC domain of ATX (and ENPP1 and ENPP3) encompasses an EF-hand-like (EFL) motif26 ( 739 DYNYDGLRDTEDE 751 ), crucial for activity for ENPP1 and ENPP331, but reportedly not for ENPP232. This 13-residue EFL motif binds Ca +2 (Supplementary Fig.…”

Section: Resultsmentioning

confidence: 99%

Structural basis of substrate discrimination and integrin binding by autotaxin

Hausmann

Kamtekar

Christodoulou

et al. 2011

Nat Struct Mol Biol

264

390

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Autotaxin (ATX) or ecto-nucleotide pyrophosphatase/phosphodiesterase-2 (ENPP2) is a secreted lysophospholipase D that generates the lipid mediator lysophosphatidic acid (LPA), a mitogen and chemo-attractant for many cell types. ATX-LPA signaling has roles in various pathologies including tumour progression and inflammation. However, the molecular basis of substrate recognition and catalysis, and the mechanism of interaction with target cells, has been elusive. Here we present the crystal structure of ATX, alone and in complex with a small-molecule inhibitor. We identify a hydrophobic lipid-binding pocket and map key residues required for catalysis and selection between nucleotide and phospholipid substrates. We show that ATX interacts with cell-surface integrins via its N-terminal somatomedin-B-like domains, using an atypical mechanism. Our results define determinants of substrate discrimination by the ENPP family, suggest how ATX promotes localized LPA signaling, and enable new approaches to target ATX with small-molecule therapeutics.

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Section: Resultsmentioning

confidence: 99%

Structural basis of substrate discrimination and integrin binding by autotaxin

Hausmann

Kamtekar

Christodoulou

et al. 2011

Nat Struct Mol Biol

264

390

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“…Gelatin‐coated Nuclepore membranes used in these modified Boyden chambers were fixed and stained using Diff‐Quik reagents (American Scientific Products). Chemotaxis was quantified densitometrically using EagleSight Software v. 3.2 (Stratagene) for data analysis as described previously [16].…”

Section: Methodsmentioning

confidence: 99%

Autotaxin promotes motility via G protein‐coupled phosphoinositide 3‐kinase γ in human melanoma cells

et al. 2002

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Autotaxin (ATX), an exo-nucleotide pyrophosphatase and phosphodiesterase, stimulates tumor cell motility at subnanomolar levels and augments invasiveness and angiogenesis. We investigated the role of G protein-coupled phosphoinositide 3-kinase Q Q (PI3KQ Q) in ATX-mediated tumor cell motility stimulation. Pretreatment of human melanoma cell line A2058 with wortmannin or LY294002 inhibited ATX-induced motility. ATX increased the PI3K activity in p110Q Q, but not p85, immunoprecipitates. This effect was abrogated by PI3K inhibitors or inhibited by pertussis toxin. Furthermore, stimulation of tumor cell motility by ATX was inhibited by catalytically inactive form of PI3KQ Q, strongly indicating the crucial role of PI3KQ Q for ATXmediated motility in human melanoma cells ß

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“…The Ca 2+ -binding site is also present in the structures of NPP1 and NPP2. This motif appears to be important for the structural integrity of the nuclease-like domain and so for the catalytic activity of NPP1 1 and NPP3 36 but not for NPP2 37 . In addition, N754 and K452 form a hydrogen bond interaction across the domain interface.…”

Section: Resultsmentioning

confidence: 98%

Crystal structure and substrate binding mode of ectonucleotide phosphodiesterase/pyrophosphatase-3 (NPP3)

Dohler

Zebisch

Sträter

2018

Sci Rep

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Ectonucleotide phosphodiesterase/pyrophosphatase-3 (NPP3) is a membrane-bound glycoprotein that regulates extracellular levels of nucleotides. NPP3 is known to contribute to the immune response on basophils by hydrolyzing ATP and to regulate the glycosyltransferase activity in Neuro2a cells. Here, we report on crystal structures of the nuclease and phosphodiesterase domains of rat NPP3 in complex with different substrates, products and substrate analogs giving insight into details of the catalytic mechanism. Complex structures with a phosphate ion, the product AMP and the substrate analog AMPNPP provide a consistent picture of the coordination of the substrate in which one zinc ion activates the threonine nucleophile whereas the other zinc ion binds the phosphate group. Co-crystal structures with the dinucleotide substrates Ap4A and UDPGlcNAc reveal a binding pocket for the larger leaving groups of these substrates. The crystal structures as well as mutational and kinetic analysis demonstrate that the larger leaving groups interact only weakly with the enzyme such that the substrate affinity is dominated by the interactions of the first nucleoside group. For this moiety, the nucleobase is stacked between Y290 and F207 and polar interactions with the protein are only formed via water molecules thus explaining the limited nucleobase selectivity.

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

Cited by 25 publications

References 21 publications

Structural basis of substrate discrimination and integrin binding by autotaxin

Structural basis of substrate discrimination and integrin binding by autotaxin

Autotaxin promotes motility via G protein‐coupled phosphoinositide 3‐kinase γ in human melanoma cells

Crystal structure and substrate binding mode of ectonucleotide phosphodiesterase/pyrophosphatase-3 (NPP3)

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