Pyrophosphates play important roles in a wide variety of biological processes. However, the intrinsic chemical and enzymatic lability of the pyrophosphate moiety complicates the study of the biological processes in which these molecules are involved. The research described in this Thesis focuses on the development of non-hydrolyzable pyrophosphate analogues as inhibitors and structural probes for investigating the structures and mechanisms of enzymes that utilize pyrophosphate-containing molecules as substrates. To facilitate these studies, new reagents and synthetic methodologies were developed, while existing methods were further optimized.
Chapter 1 introduces a variety of biologically relevant pyrophosphate-containing molecules, with particular emphasis on cytidine diphosphate glycerol (CDP-Gro) and cytidine diphosphate ribitol (CDP-Rbo), which are involved in wall teichoic acid (WTA) biosynthesis, as well as nucleoside diphosphate (NDP) sugars, which...
Show morePyrophosphates play important roles in a wide variety of biological processes. However, the intrinsic chemical and enzymatic lability of the pyrophosphate moiety complicates the study of the biological processes in which these molecules are involved. The research described in this Thesis focuses on the development of non-hydrolyzable pyrophosphate analogues as inhibitors and structural probes for investigating the structures and mechanisms of enzymes that utilize pyrophosphate-containing molecules as substrates. To facilitate these studies, new reagents and synthetic methodologies were developed, while existing methods were further optimized.
Chapter 1 introduces a variety of biologically relevant pyrophosphate-containing molecules, with particular emphasis on cytidine diphosphate glycerol (CDP-Gro) and cytidine diphosphate ribitol (CDP-Rbo), which are involved in wall teichoic acid (WTA) biosynthesis, as well as nucleoside diphosphate (NDP) sugars, which serve as donor substrates for glycosyltransferases. The chapter also provides an overview of strategies for generating pyrophosphate mimics, focusing on the synthesis of bisphosphonate and oxy-sulfonyl carbamate analogues, thereby providing the foundation for the design and synthesis of stable analogues described in the subsequent chapters.
Chapter 2 describes the design and synthesis of stabilized CDP-glycerol and CDP-ribitol analogues incorporating O- or N-sulfonyl carbamate and phosphoryl carbamate groups as potential inhibitors of enzymes involved in WTA biosynthesis.
Chapter 3 describes the preparation of two UDP-GlcNAc analogues as glycosyltransferase inhibitors using a phosphoramidite coupling strategy and a newly developed orthogonally protected phosphonylmethylphosphonate reagent. The reagent and complementary methodology were subsequently applied in Chapter 5.
Chapter 4 describes an efficient orthogonal deprotection–condensation strategy for the stepwise functionalization and coupling of primary alcohols with a symmetrical difluoromethylene bisphosphonate reagent. The key reagent could be prepared on a large scale, providing sufficient quantities for the synthesis of various unsymmetrical difluoromethylene bisphosphonate diesters. The versatility of this strategy was demonstrated through the construction of a series of pyrophosphate mimics. The methodology was subsequently extended to the synthesis of UDP-galactose analogues in Chapter 5.
Chapter 5 focuses first on the synthesis of carbagalactose, the building block required for the preparation of UDP-galactose analogues. The phosphonylmethylphosphonate reagents developed in Chapter 3 were then employed for the synthesis of P–CH₂–P UDP-carbagalactose. Further optimization of the methodology described in Chapter 4 enabled its extension to the coupling of secondary alcohols, allowing access to P–CF₂–P UDP-carbagalactose. The inhibitory activities of these analogues against human α-1,4-galactosyltransferase (A4GALT) were subsequently evaluated.
Chapter 6 summarizes the research described in this Thesis and outlines future research directions based on the design of pyrophosphate mimics and the synthetic methodologies developed throughout this work.
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