Mechanistic studies on chemical instabilities of recombinant proteins

Protein molecules are being widely used as pharmaceuticals for treating diseases ranging from diabetes and haemophilia to various types of cancers due to their great potency and specificity. However, these macromolecules are intrinsically unstable in aqueous solutions, due to the existence of various physical and chemical degradation pathways. Degraded protein molecules have much reduced biological functions, and may also have adverse effects such as immunogenriicity or pharmacokinetic issues. Thus, understanding the underlying mechanisms of these degradation pathways is essential for rationally devising better ways to stabilize protein pharmaceuticals and extends their applicability. In this thesis, two important types of chemical degradation pathways, the oxidation of methionine residues and the hydrolysis of peptide bonds in monoclonal antibody molecules, are investigated from a mechanistic point of view. In the first half of the thesis, oxidation 'of methionine residues in a model protein G-CSF (Granulocyte-Colony Stimulating Factor) was studied to address the issue of how protein structure affects its reactivity. Comparative oxidation studies were performed where the kinetics of oxidation of methionine residues by hydrogen peroxide (H₂0₂) in G-CSF and corresponding chemically synthesized peptides thereof were measured at different temperatures. To assess structural effects, equilibrium denaturation experiments also were conducted on G-CSF to obtain the free energy of unfolding as a function of temperature.

Department dc:contributor.department

Massachusetts Institute of Technology. Dept. of Chemical Engineering.

Grantor dc:publisher

Massachusetts Institute of Technology

Year dc:date.issued

2009

Author dc:creator

• Pan, Bin, Ph. D. Massachusetts Institute of Technology

Advisor dc:contributor.advisor

• Daniel I. C. Wang and Bernhardt L. Trout.

• Chemical Engineering.

Statement dc:rights

• M.I.T. theses are protected by copyright. They may be viewed from this source for any purpose, but reproduction or distribution in any format is prohibited without written permission. See provided URL for inquiries about permission.

Licence dc:rights.uri

• http://dspace.mit.edu/handle/1721.1/7582

Language dc:language.iso

eng

Handle dc:identifier.uri

http://hdl.handle.net/1721.1/51629

OAI identifier oai:identifier

oai:dspace.mit.edu:1721.1/51629