Abstract
Endostatin, a C-terminal fragment of collagen XVIII, binds to TG-2 (transglutaminase-2) in a cation-dependent manner. Recombinant human endostatin binds to TG-2 with an affinity in the nanomolar range (Kd=6.8 nM). Enzymatic assays indicated that, in contrast with other extracellular matrix proteins, endostatin is not a glutaminyl substrate of TG-2 and is not cross-linked to itself by the enzyme. Two arginine residues of endostatin, Arg27 and Arg139, are crucial for its binding to TG-2. They are also involved in the binding to heparin [Sasaki, Larsson, Kreuger, Salmivirta, Claesson-Welsh, Lindahl, Hohenester and Timpl (1999) EMBO J. 18, 6240–6248], and to α5β1 and αvβ3 integrins [Faye, Moreau, Chautard, Jetne, Fukai, Ruggiero, Humphries, Olsen and Ricard-Blum (2009) J. Biol. Chem. 284, 22029–22040], suggesting that endostatin is not able to interact simultaneously with TG-2 and heparan sulfate, or with TG-2 and integrins. Inhibition experiments support the hypothesis that the GTP-binding site of TG-2 is a potential binding site for endostatin. Endostatin and TG-2 are co-localized in the extracellular matrix secreted by endothelial cells under hypoxia, which stimulates angiogenesis. This interaction, occurring in a cellular context, might participate in the concerted regulation of angiogenesis and tumorigenesis by the two proteins.
- endostatin (ES)
- extracellular matrix
- protein–protein interaction
- surface plasmon resonance (SPR) binding assays
- transglutaminase-2 (TG-2)
Abbreviations: COL18A1, collagen XVIII, α1; EBNA, Epstein–Barr virus nuclear antigen; ERK, extracellular-signal-regulated kinase; FAK, focal adhesion kinase; HEK, human embryonic kidney; HIF, hypoxia-inducible factor; HUVEC, human umbilical vein endothelial cell; MAPK, mitogen-activated protein kinase; MMP, matrix metalloproteinase; MT1, membrane-type 1; NC1 domain, entire C-terminal globular domain of collagen XVIII; SPR, surface plasmon resonance; TBS, Tris-buffered-saline; TG-2, transglutaminase-2; TLR, Toll-like receptor; VEGF, vascular endothelial growth factor; VEGFR, VEGF receptor
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