Spontaneous formation of L-isoaspartate and gain of function in fibronectin.
Isoaspartate formation in extracellular matrix proteins, by aspartate isomerization or asparagine deamidation, is generally viewed as a degradation reaction occurring in vivo during tissue aging. For instance, non-enzymatic isoaspartate formation at RGD-integrin binding sites causes loss of cell adhesion sites, which in turn can be enzymatically "repaired" to RGD by protein-l-isoAsp-O-methyltransferase. We show here that isoaspartate formation is also a mechanism for extracellular matrix activation. In particular, we show that deamidation of Asn263 at the Asn-Gly-Arg (NGR) site in fibronectin N-terminal region generates an alpha(v)beta3-integrin binding site containing the L-isoDGR sequence, which is enzymatically "deactivated" to DGR by protein-L-isoAsp-O-methyltransferase. Furthermore, rapid NGR-to-isoDGR sequence transition in fibronectin fragments generates alpha(v)beta3 antagonists (named "isonectins") that competitively bind RGD binding sites and inhibit endothelial cell adhesion, proliferation, and tumor growth. Time-dependent generation of isoDGR may represent a sort of molecular clock for activating latent integrin binding sites in proteins.
Curnis
Flavio
F
Department of Oncology, Cancer Immunotherapy and Gene Therapy Program and Italian Institute of Technology Network Research Unit of Molecular Neuroscience, San Raffaele Scientific Institute, Via Olgettina 58, 20132 Milan, Italy.
Longhi
Renato
R
Crippa
Luca
L
Cattaneo
Angela
A
Dondossola
Eleonora
E
Bachi
Angela
A
Corti
Angelo
A
D000595:Amino Acid Sequence; D000818:Animals; D001667:Binding, Competitive; D002448:Cell Adhesion; D005109:Extracellular Matrix / Q000378:metabolism; D005353:Fibronectins / Q000737:chemistry*; D006801:Humans; D016023:Integrins / Q000737:chemistry; D026581:Isoaspartic Acid / Q000737:chemistry / Q000502:physiology*; D008546:Melanoma, Experimental; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry
D005353:Fibronectins; D016023:Integrins; D026581:Isoaspartic Acid; D009842:Oligopeptides; D011994:Recombinant Proteins; C047981:arginyl-glycyl-aspartic acid