Current Organic Chemistry - Volume 5, Issue 8, 2001

Nucleic acids exist in a number of structural states that may comprise one to four strands. Common features involve base-stacking and pairing of complementary bases via H-bonded interactions, although a wide variety of pairing motifs are found in higher-order structures. Another property is the marked flexibility of nucleic acids reflected in their solution behaviour, thermodynamic stability and interactions with proteins and small ligand molecules. The electrostatic properties of a nucleic acid and its ability to pack into high-order structure(s) are largely determined by the anionic phosphodiester backbone. This feature is particularly evident for triple-helical assemblies, where there is close proximity between phosphates and intrinsic thermodynamic stability is strongly influenced by ionic strength. Structural stabilisation of parallel triplexes can also be modulated by the presence of positive charge, achieved through either N3-protonation of cytosine bases or the deliberate incorporation of charged residues into the third strand. Salt effects are also pronounced in four-stranded structures such as the i-motif and G-tetraplex (quadruplex) assemblies indeed, different structures are produced for the latter depending on the nature of the associated cation. High-order nucleic acid structures are currently a focus of intense interest in antisense and antigene strategies toward novel chemotherapeutic agents. An understanding of both their structural details and solution properties is essential for a rational approach to DNA-targeted drug design.

DNA polymerase I enzymes have served as model systems to study the mechanism of template-directed DNA polymerization. This process requires that the enzyme cycles through a series of conformational changes, each cycle leading to the incorporation of a nucleotide to the primer strand of the DNA. The kinetics of nucleotide incorporation has been extensively studied leading to the definition of specific steps along the cycle. Efforts to visualize these steps using X-ray crystallography have recently come to fruition, notably for one particular DNA polymerase I system, that of Klentaq1. This review focuses on the structural characterization of the various steps along the nucleotide incorporation pathway.

The replication reaction provides the molecular source for mutation, selection and evolution in biology. This reaction also requires two additional stages, transcription and translation, functioning cooperatively, to construct the materials necessary to transfer the genome accurately to progeny. These three stages together constitute biologys Central Dogma, a term that highlights their presence in all autonomous replicating organisms. The template-directed synthesis that underpins each of these stages is dependent on molecular complementarity and the energies of self-assembly, suggesting it might now be possible to borrow from the structural tool set of biology to explore, diversify and even extend these reactions into different structural skeletons. Here we demonstrate, using simple ligation reactions, that it is conceptually possible to simplify the Central Dogma to a two-stage DNA replication process. Through this system DNA sequence information can be read into different molecular skeletons, skeletons with unique catalytic properties, that when extended, are amenable to mutation, selection, and evolution of desired function.

Gene expression is a central phenomenon in all organisms. The first step in the expression of a gene is the transcription of the DNA to generate a messenger RNA, which is subsequently translated to a protein. In response to cellular and environmental cues regulatory proteins, known as transcriptional activators or repressors, either enhance or inhibit transcription of specific genes. Here, a summary of our current understanding of this initial step in gene expression is presented, along with the recent advances in creating synthetic analogs of regulatory transcription factors. The study of transcriptional regulation of an individual gene as well as that of all genes in the genome is now accessible to rational chemical intervention. The current challenges in the design of small-molecule transcription factors and their potential role in generating novel transcription-based therapeutics are also discussed.