Chemical Protein Synthesis - my Journey from SPPS to Chemical Ligation
Stephen Kent
Professor, University of Chicago
The total synthesis of proteins was one of the 'Grand Challenges' of 20th century chemistry. Despite decades of development by skilled organic chemists throughout the world, conventional synthetic methods and stepwise SPPS were able to make only the smallest proteins. In 1992 we introduced the chemical ligation principle - the condensation of unprotected peptide segments by chemoselective reaction enabled by formation of an unnatural bond at the ligation site. [1] In 1994 we introduced the native chemical ligation reaction, thiol ester-mediated condensation of unprotected peptide segments to give a native peptide bond at the ligation site. [2] In combination with peptide synthesis by state- of-the-art SPPS, modern chemical ligation methods enable total chemical synthesis of proteins including enzymes. [3,4] Synthetic protein products are characterized by high resolution methods including LCMS, multidimensional NMR, and X-ray crystallography. Chemical protein synthesis gives precise, atom-
by-atom control over the covalent structure of a protein molecule. Case studies of the application of chemical protein synthesis to the enzyme HIV-1 protease will be described. Efficient total synthesis of human insulin will also be presented, together with site-specific 13 C= 18 O isotope labelling of individual peptide bonds in the insulin molecule for time-resolved FTIR spectroscopy.
References
1. Constructing proteins by dovetailing unprotected synthetic peptides: backbone engineered
HIV protease. M. Schnölzer, S. Kent Science, 256, 221-225 (1992).
2. Synthesis of proteins by native chemical ligation. Philip E. Dawson, Tom W. Muir, Ian Clark-
Lewis, Stephen B.H. Kent, Science, 266, 776-779 (1994).
3. Chemical protein synthesis: inventing synthetic methods to decipher how proteins work.
Stephen B.H. Kent, Bio Org Med Chem., 25, 4926-4937 (2017).
4. Novel protein science enabled by total chemical synthesis. Kent SBH. Protein Science 2019;
28:313–328
Stephen B. H. Kent received his first two university degrees in his native New Zealand and his Ph.D. (Organic Chemistry) from University of California, Berkeley. Following postdoctoral work with Bruce Merrifield, Professor Kent held faculty appointments at the Rockefeller University, the California Institute of Technology, and The Scripps Research Institute. He was Founder of Gryphon Sciences and served as its Chief Scientific Officer from 1997-2000. In 2001, he joined the faculty of the University of Chicago, where since June 2021 he is Professor Emeritus of Chemistry, Biochemistry and Molecular Biology.
Over the past several decades, Stephen Kent has profoundly shaped the field of peptide and protein chemistry. His many and diverse contributions have altered the way scientists tackle the study of these macromolecules. He pioneered the total chemical synthesis of protein molecules and showed it to be a practical and versatile method for the production of large quantities of high purity material for detailed mechanistic and structural investigations. In particular, the Kent chemical ligation concept together with the native chemical ligation reaction made proteins and enzymes accessible to direct chemical investigation via convergent, chemoselective assembly of unprotected synthetic peptide segments.
Since its introduction in 1994, native chemical ligation has become robust, reliable, and widely used. Efficient total chemical synthesis of a wide range of protein molecules, including the enzymes HIV-1 protease and human lysozyme, the protein hormone human insulin, glycoprotein mimetics of erythropoietin, together with numerous mirror image protein molecules for his pioneering studies of racemic protein crystallography, highlight the practical utility of such approaches. As these examples attest, Stephen Kent has had a gift, throughout his career, for developing new chemistries and for utilizing them to reveal the molecular basis of protein function.
His exceptional achievements in scientific research have previously been recognized by the receipt of major distinctions, including the ACS Hirschmann Award in Peptide Chemistry (1994), the Protein Society E.T. Kaiser Award (2002), the Vincent du Vigneaud (2004) and R. Bruce Merrifield (2009) Awards of the American Peptide Society, the European Peptide Society Rudinger Medal (2010), the Japanese Peptide Society Akabori Medal (2010), the ACS Alfred Bader Award in Bioorganic Chemistry (2011), and more.