Specifically targeted catalytic antagonists and uses thereof
This invention provides chimeric molecules that are catalytic antagonists of a target molecule. The catalytic antagonists of this invention preferably comprise a targeting moiety attached to an enzyme that degrades the molecule specifically bound by the targeting moiety. The catalytic antagonists of this invention thus bind to a target recognized by the targeting moiety (e.g., a receptor) the enzyme component of the chimera then degrades all or part of the target. This typically results in a reduction or loss of activity of the target and release of the chimeric molecule. The chimeric molecule is then free to attack and degrade another target molecule.
1. A method of degrading a target molecule, said method comprising contacting said target molecule with a catalytic antagonist comprising a carbohydrate targeting moiety attached to a subtilisin-type serine protease,
wherein said targeting moiety specifically binds to said target molecule and said protease degrades said target molecule, resulting in the release of said antagonist, thereby allowing said antagonist to bind and degrade another target molecule.
2. The method of claim 1 , wherein said targeting moiety is joined to said serine protease through the sulfur group on a cysteine.
3. The method of claim 2 , wherein said cysteine is a cysteine that is introduced into said serine protease by substituting a native amino acid other than cysteine with said cysteine.
4. The method of claim 3 , wherein said cysteine is a cysteine that is substituted for a native amino acid other than cysteine in or near a subsite comprising a substrate binding site of said serine protease.
5. The method of claim 4 , wherein said cysteine is a subsite selected from the group consisting of an S1 subsite, an S1′ subsite, and an S2 subsite.
6. The method of claim 5 , wherein said serine protease is a Bacillus lentos subtilisin.
7. The method of claim 5 , wherein said cysteine is substituted for an amino acid corresponding to a reference residue in a Bacillus lentos subtilisin, where said reference residue is at a residue selected from the group consisting of residue 156, residue 166, residue 217, residue 222, residue 62, residue 96, residue 104, residue 107, residue 189, and residue 209.
8. The method of claim 1 , wherein said target is a molecule present on the surface of a cell.
9. The method of claim 8 , wherein said molecule present on the surface of a cell is a molecule forming a receptor.
10. The method of claim 8 , wherein said molecule present on the surface of a cell is a ligand.
11. The method of claim 8 , wherein said molecule present on the surface of a cell is component of a cell wall.
12. The method of claim 8 , wherein said molecule present on the surface of a cell is component of a cell membrane.
13. The method of claim 1 , wherein said targeting moiety is -thioethyl D-mannopyranoside.