Composition exhibiting a von Willebrand Factor (vWF) protease activity with the amino acid sequence AAGGILHLELLV
The invention relates to vWF cleaving entities having a molecular weight of 180 kD, 170 kD, 160 kD, 120 kD or 110 kD and an N-terminal amino acid sequence of AAGGILHLELLV, vWF cleaving complexes and methods for their production.
1. A method for purification of a vWF protease polypeptide comprising the steps of:
(a) contacting a solution comprising vWF protease polypeptide with an antibody that: binds to a polypeptide present in human plasma that exhibits vWF protease activity, has a molecular weight from about 180 kD to about 120 kD as determined by SDS-PAGE under reducing conditions, and comprises the amino acid sequence AAGGILHLELLV (SEQ ID NO: 1);
wherein the polypeptide is contacted with the antibody under physiological conditions whereby the polypeptide binds to the antibody,
(b) eluting the vWF protease polypeptide from the antibody.
2. The method according to claim 1 , wherein the antibody is immobilized on an affinity chromatography gel.
3. The method according to claim 1 , wherein the solution is human plasma.
4. The method according to claim 1 , wherein the antibody binds to the amino acid sequence
(SEQ ID NO:4)
AGGILHLELLVAVGPDVFQAHQEDTERYVLTNLNIGAELLRDPSLGAQFR
VHLVKMVILTEPEGAPNITANLTSSLLSVCGWSQTINPEDDTDPGHADLV
LYITRFDLELPDGNRQVRGVTQLGGACSPTWSCLITEDTGFDLGVTI.
5. The method according to claim 1 , wherein the antibody is a monoclonal antibody.
6. The method according to claim 1 , further comprising a step of gel filtration or ion exchange chromatography.
7. The method of claim 6 , wherein the ion exchange chromatography step is anion exchange.
8. The method according to claim 1 , further comprising a gel filtration step and an ion exchange chromatography step.
9. The method according to claim 8 , wherein the ion exchange chromatography is anion exchange.