Protein folding

• US 7,893,221 B2
• Filed: 03/09/2007
• Issued: 02/22/2011
• Est. Priority Date: 03/11/2006
• Status: Expired due to Fees

• Current Assignee: Renovo Ltd. (Bentley Park Ltd.)
• Original Assignee: Renovo Ltd. (Bentley Park Ltd.)
• Inventors: Higginbottom, Simon, Ferguson, Mark William James, Occleston, Nick, Mellors, Phillip, O'Kane, Sharon, Laverty, Hugh Gerard
• Primary Examiner(s): Nashed; Nashaat T
• Application Number:
  US12/282,456
• Publication Number:
  US 20090181430A1
• Time in Patent Office: 1,446 Days
• Field of Search: None
• US Class Current: 530/402
• CPC Class Codes: A61P 1/16 for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics
• CPC Class Codes: A61P 17/00 Drugs for dermatological disorders
• CPC Class Codes: A61P 17/02 for treating wounds, ulcers, burns, scars, keloids, or the like
• CPC Class Codes: A61P 19/00 Drugs for skeletal disorders
• CPC Class Codes: A61P 27/02 Ophthalmic agents
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 9/00 Drugs for disorders of the cardiovascular system
• CPC Class Codes: C07K 1/1133 by redox-reactions involving cystein/cystin side chains
• CPC Class Codes: C07K 14/495 Transforming growth factor [TGF]

Abstract

The present invention concerns a method for folding a Transforming Growth Factor Beta, or a functional analogue thereof, into a dimeric, biologically active form. The method involves adding solubilized, unfolded monomeric growth factor to a solution containing 2-(cyclohexylamino)-ethanesulfonic acid (CHES) or a functional analogue thereof and a low molecular weight sulfhydryl/disulfide redox system. The solution is then incubated under conditions suitable for generating dimeric biologically active Transforming Growth Factor Beta.

Claims

1. A method of forming a dimeric, biologically active transforming growth factor beta 3 comprising adding solubilized, unfolded monomeric transforming growth factor beta 3 to a solution comprising:

(i) 0.7 M 2-(cyclohexylamino)-ethanesulfonic acid (CHES);

(ii) a low molecular weight sulfhydryl/disulfide redox system comprising about 2 mM reduced glutathione and about 0.4 mM oxidized glutathione; and

(iii) 1 M NaCl,

wherein the concentration of transforming growth factor beta 3 in the solution is about 0.25mg/mL and the solution is at a pH of about 9.5,

and incubating the unfolded monomeric transforming growth factor beta 3 in the solution until a dimeric biologically active transforming growth factor beta 3 is formed, wherein the forming of dimeric biologically active transforming growth factor beta 3 in the solution is completed within about 2 days.