Therapeutic Proteins: Strategies in Synthetic Biology

Seminar

Title: Therapeutic Proteins: Strategies in Synthetic Biology

Speaker: Gonçalo Bernardes

Affiliation: ETH Zürich, Switzerland

Host: Inês Cardoso Pereira

Abstract:

Post-translational modification (PTM) is an important step in protein biosynthesis that increases the range of functions of a protein. Selective engineering of proteins using PTM has been used to develop therapeutic proteins for the treatment of cancer, cystic fibrosis, diabetes, anaemia, and more. The market for therapeutic proteins is large ($37 billion in 2003) and growing (>15% growth p.a.), and is expected to reach $90 billion by 2010. However, the accurate and consistent bioprocessing of therapeutic proteins is very difficult, due to their sensitivity to preparative conditions. This presents challenges for the manufacture, regulation and safety of therapeutic proteins. There is an immediate need for methods that enable specific and controllable modification of proteins.

Site-selective chemical modification has emerged as a powerful strategy for modulating macromolecular function.[1] Precise chemical modification of proteins is a daunting challenge in chemoselectivity: the reaction must proceed efficiently in buffered aqueous media at or near room temperature and select a single reactive residue among hundreds of competing functional groups. In the work presented, the unique reactivity of cysteine is explored. For cysteine, the nucleophilicity of the side chain thiol is harnessed in selective formation of thioethers. Two complimentary methods have been developed: The first is a reductive conversion of disulfides to thioethers mediated by electron rich phosphines[2] and the second is an oxidative route that hinges on a novel, efficient conversion of cysteine to dehydroalanine which can then be trapped with thiol nucleophiles.[3] Additionally, chemical introduction of S-ally cysteine allowed us to take advantage of the enhanced reactivity of allyl sulfides in CM, and we were able to post-translationally modify proteins via carbon-carbon bond formation.[4] This work establishes a new standard in substrate sensitivity and complexity in olefin metathesis in water, which current benchmark for homogeneous aqueous CM was so far the self-metathesis of simple unsaturated alcohols such as allyl alcohol. Finally, zinc-mediated addition of alkyl iodides to dehydroalanine containing proteins allows creation of more than 30 protein mutants from a single round of expression. These methods are efficient strategies to access phosphorylated, glycosylated, polyprenylated, lysine-methylated proteins, and natural and unnatural amino acids that so far were impossible to incorporate into proteins.

[1] Davis, B. G. Science, 2004, 303, 480-482.
[2] Bernardes, G.J.L.; Grayson, E.J.; Thompson, S.; Chalker, J.M.; Errey, J.C.; El Oualid, F.; Claridge, T.D.W.; Davis, B.G. Angew. Chem. Int. Ed. 2008, 47, 2244-2247.
[3] Bernardes, G.J.L.; Chalker, J.M.; Errey, J.C.; Davis, B.G. J. Am. Chem. Soc. 2008, 130, 5052-5053. Article highlighted in C&E news March 31, 2008 and Nature Chem. Biol. 2008, 4, 527-528.
[4] Lin, A.Y.; Chalker, J.M.; Floyd, N.; Bernardes, G.J.L.; Davis, B.G. J. Am. Chem. Soc. 2008, 130, 9642-9643. Article highlighted in C&E news July 14, 2008 and Nature Chem. Biol. 2008, 4, 527-528.

EDUCATION

            Ph.D. in Organic Chemistry, Sept 2004 - May 2008

            University of Oxford, Oxford, UK

            Thesis advisor: Prof. Benjamin G. Davis

            Thesis Committee: Prof. Carolyn R. Bertozzi (Berkeley, CA) and Prof. Rob Field (Norwich, UK)         

            Honours Degree in Chemistry, final classification of 16 (sixteen), 1999 - Jul 2004

            University of Lisbon, Faculty of Sciences, Lisbon, Portugal                           

EXPERIENCE

2008-present Postdoctoral Research Fellow in Carbohydrate Chemistry and Chemical Biology

            Swiss Federal Institute of Tecnhology (ETH), Zürich

            Mentor: Prof. Peter H. Seeberger

2008-present Academic Visitor Department of Chemistry, University of Oxford

2008 Postdoctoral Research Fellow in Chemical Biology and Chemical Medicine

            University of Oxford, International AIDS Vaccine Initiative

            Mentor: Prof. Benjamin G. Davis

2006-2008 Class Tutor in Biological Chemistry, Dept of Biochemistry, University of Oxford

2003-2004 Research Training at Glycoform Ldt, Oxford, UK