The de novo chemical synthesis of proteins has the potential to rapidly accelerate the study of proteins by providing rapid access to natural and rationally designed unnatural proteins. Currently, however, chemical synthesis of proteins is limited to relatively small sized polypeptides and glycopeptides, due to a number of practical factors. Generally, the efficiency of ligation of peptide fragments decreases with increasing peptide length, where peptide aggregation is an often observed obstacle. We propose an alternative paradigm for the protection of peptides against aggregation and the size limitations of native ligation, in contrast to the introduction of extraneous protection functionalities and auxiliaries. The induction of secondary structure in solution for peptide fragments that are predisposed to the formation of alpha-helices should serve to protect the peptides against destructive aggregation by rigidifying the peptide backbone into a compact conformation. This line of reasoning is in stark contrast to the typical native chemical ligation protocol that is historically performed exclusively under denaturing conditions. In addition to reduced propensity towards aggregation, we expect the helical protein fragments to have increased efficiency of ligation due to the conformation compactness and rigidity, and a corresponding reduction in the negative correlation between peptide complexity and ligation efficiency. In order to investigate this approach towards peptide ligation, the bromodomain of human protein ATAD2 will be synthesized. In only the past few years, this protein has been identified as upregulated in breast, prostate, lung, and ovarian tumors, and numerous studies have correlated the over expression of ATAD2 with cancer growth and patient prognosis. Considering the involvement of ATAD2 with protein transcription and mitosis along with the upregulation observed in a wide variety of tumor types, it is no surprise that ATAD2 has been identified as a potential therapeutic target. The bromodomain of ATAD2 is an ideal target to test the helical stabilization hypothesis because it is made up of five alpha-helices, each of ideal size for solid phase peptide synthesis, and includes two relatively hydrophobic regions with potential for aggregation. This investigation has the potential for broad implications, as the strategic revision for the chemical synthesis of proteins that is proposed applies new rules for the disconnection of polypeptides into their corresponding fragments.

Public Health Relevance

The de novo chemical synthesis of proteins is currently lacking an organized strategy for peptide disconnections, which is preventing widespread adoption. A strategy for protein disconnection based on secondary structure and helical content has been proposed, and has broad implications for the rapid synthesis and investigation of natural and synthetic proteins. Synthesis of the bromodomain region of human ATAD2, a protein associated with multiple forms of aggressive tumor growth and whose upregulation has been correlated with poor patient prognosis, serves as an ideal target to explore the relationship between induced helical character and synthetic strategies.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Postdoctoral Individual National Research Service Award (F32)
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Special Emphasis Panel (ZRG1-F04-A (20))
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Barski, Oleg
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Sloan-Kettering Institute for Cancer Research
New York
United States
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Levinson, Adam M; McGee, John H; Roberts, Andrew G et al. (2017) Total Chemical Synthesis and Folding of All-l and All-d Variants of Oncogenic KRas(G12V). J Am Chem Soc 139:7632-7639
Creech, Gardner S; Paresi, Chelsea; Li, Yue-Ming et al. (2014) Chemical synthesis of the ATAD2 bromodomain. Proc Natl Acad Sci U S A 111:2891-6