Strategies for the Synthesis of Sulfopeptides and Sulfoproteins as Therapeutic Leads

Abstract

Tyrosine sulfation is a common post-translational modification (PTM) in multicellular organisms and implicated in numerous physiological and pathological processes occurring in animals. However, the understanding and study of this PTM is hampered by the limited access to sulfoproteins through conventional biotechnology methods. Chemical synthesis of post-translationally modified proteins provides a powerful way to access homogenous variants of sulfated peptides and proteins. This thesis describes the development of new synthetic methodologies and biological studies to interrogate the functional role of the PTM. Chapter 2 describes the convergent syntheses of two anti-thrombotic proteins and their sulfated variants, namely chimadanin and the Anopheles albimanus protein through the use of trifluoroethanthiol (TFET)-mediated ligation–desulfurisation chemistry. These synthetic (sulfo)forms of these proteins enabled unequivocal determination of the functional role of tyrosine sulfation on thrombin inhibition. Subsequent in vitro and in vivo studies have revealed that tyrosine sulfation tremendously increases thrombin inhibitory activity of these thrombin inhibitors. Chapter 3 describes the studies toward the synthesis of evasin-4, a small chemokine-binding protein (CKBP). This chapter provides useful insights into the strength and limitations of several protein ligation techniques. In particular, the additive-free selenoester-selenocystine ligation chemistry applied in the synthesis can greatly expand the scope of peptide ligation methodologies and provides multiple levels of orthogonality into the ligation-based assembly of polypeptides. Chapter 4 describes an efficient approach to the synthesis of sulfopeptide library through orthogonal-protecting-group and solid-phase sulfation strategies. We were able to access a library of eight (sulfo)forms of an N-terminal fragment of the HIV-1 coreceptor CC chemokine receptor 5 for detailed anti-HIV testing.