Cloning, expression and functional characterization of the full-length murine ADAMTS13.
Functional deficiency or absence of the human von Willebrand factor (VWF)-cleaving protease (VWF-cp), recently termed ADAMTS13, has been shown to cause acquired and congenital thrombotic thrombocytopenic purpura (TTP), respectively. As a first step towards developing a small animal model of TTP, we have cloned the complete (non-truncated) murine Adamts13 gene from BALB/c mice liver poly A+ mRNA. Murine ADAMTS13 is a 1426-amino-acid protein with a high homology and similar structural organization to the human ortholog. Transient expression of the murine Adamts13 cDNA in HEK 293 cells yielded a protein with a molecular weight of approximately 180 kDa which degraded recombinant murine VWF (rVWF) in a dose-dependent manner. The cleavage products of murine rVWF had the expected size of 140 and 170 kDa. Murine ADAMTS13 was inhibited by EDTA and the plasma from a TTP patient.
Bruno
K
K
Baxter BioScience, Vienna, Austria.
Völkel
D
D
Plaimauer
B
B
Antoine
G
G
Pable
S
S
Motto
D G
DG
Lemmerhirt
H L
HL
Dorner
F
F
Zimmermann
K
K
Scheiflinger
F
F
D051722:ADAM Proteins; D000071120:ADAMTS13 Protein; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry; D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D002460:Cell Line; D003001:Cloning, Molecular; D017931:DNA Primers / Q000737:chemistry; D018076:DNA, Complementary / Q000378:metabolism; D030541:Databases, Genetic; D004305:Dose-Response Relationship, Drug; D004492:Edetic Acid / Q000494:pharmacology; D006801:Humans; D008666:Metalloendopeptidases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D008957:Models, Genetic; D008969:Molecular Sequence Data; D010802:Phylogeny; D011061:Poly A / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011697:Purpura, Thrombotic Thrombocytopenic / Q000097:blood; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D014018:Tissue Distribution; D014162:Transfection; D014841:von Willebrand Factor / Q000378:metabolism
D000596:Amino Acids; D017931:DNA Primers; D018076:DNA, Complementary; D012333:RNA, Messenger; D011994:Recombinant Proteins; D014841:von Willebrand Factor; D011061:Poly A; D012313:RNA; D004492:Edetic Acid; D051722:ADAM Proteins; C502939:ADAMTS13 protein, mouse; D008666:Metalloendopeptidases; D000071120:ADAMTS13 Protein; C099604:ADAMTS13 protein, human