Affinity tags and processes for purifying and immobilizing proteins using same
The present disclosure provides affinity tags, fusion proteins comprising one or more affinity tags, compositions comprising a fusion protein, methods of purifying a protein using an affinity tag, and devices for purifying a protein using an affinity tag.
1. A protein comprising a sequence having at least 75% sequence identity to SEQ. ID. NO: 1, wherein the protein is bound to a silicon oxide substrate.
2. A substrate comprising a protein bound to a silicon oxide substrate, the protein comprising an amino acid sequence having at least 75% sequence identity to the amino acid sequence of SEQ ID NO: 1.
3. The substrate of claim 2 wherein the bound protein has an amino acid sequence having at least 80%, at least 90% or at least 95% sequence identity to the amino acid sequence of SEQ. ID. NO: 1.
4. The substrate of claim 2 , wherein the bound protein is a portion of a fusion protein.
5. The substrate of claim 4 , wherein the fusion protein comprises a polyclonal antibody, a monoclonal antibody, a complement determining region-grafted antibody preparation, a hybrid antibody, an altered antibody, a F(ab)′ 2 fragment, a Fab molecule, a Fv fragment, a single domain antibody, a chimeric antibody, or a fragment thereof.
6. The substrate of claim 2 , wherein the substrate comprises silica.
7. A kit for purifying a protein comprising an amino acid sequence having at least 75% sequence identity to the amino acid sequence of SEQ. ID. NO:1, the kit comprising;
a silicon oxide substrate that binds to the protein;
a vessel for retainably holding the silicon oxide substrate;
a receptacle for receiving eluent passing through the vessel; and
a releasing agent.
8. The kit of claim 7 further comprising instructions for using the kit to purify the protein.
9. The kit of claim 7 , wherein the releasing agent comprises arginine and/or lysine.
10. The kit of claim 7 , wherein the releasing agent comprises a protease.
11. A method of isolating a protein comprising an amino acid sequence having at least 75% sequence identity to the amino acid sequence of SEQ. ID. NO: 1, the method comprising:
contacting a silicon oxide substrate with a fluid comprising the protein for a time sufficient to allow the protein to bind to the silicon oxide substrate;
contacting the silicon oxide substrate with a washing fluid; and
contacting the silicon oxide substrate with a releasing agent to release the bound protein.
12. The method of claim 11 , wherein the releasing agent comprises lysine and/or arginine.
13. The method of claim 11 , wherein the releasing agent comprises a protease.
14. The method of claim 11 , wherein the silicon oxide substrate comprises silica, borosilicate, controlled pore glass, quartz, and/or oxidized silicon.
15. The method of claim 11 , further comprising contacting the released protein with a protease to cleave the SEQ ID NO.: 1 from the released protein.