Harding Group, School of Chemistry, UNSW

The Harding Group - Research

Antifreeze Proteins and Glycoproteins

Many Antarctic and Arctic fish contain antifreeze proteins (AFPs) and glycoproteins (AFGPs) which prevent ice crystal growth down to -1.9 °C thus allowing the fish to survive in sub-zero waters. These biological antifreezes have tremendous medical and industrial applications where low temperature storage is required and ice crystallization is damaging e.g., including improved protection of blood platelets and human organs at low temperatures, and in by improving the smooth texture of frozen foods. The overall aim of this research is to gain an understanding of the mechanism of interaction of different classes of antifreeze proteins and antifreeze glycoproteins with the ice/water interface in order to allow rational design of synthetic proteins that are cheaper and more active than proteins from natural sources. These new antifreezes can be produced using recombinant techniques or by chemical synthesis.

Current research is focused on:
(i) the use of chemoenzymatic methods to produce AFGP mimics, that can be routinely and conveniently prepared and thus address the major limitation that has prevented commercial applications of natural AFGPs, which must be harvested from fish from Polar oceans, and contain mixtures of oligomers with structural variations depending on the exact species of fish and
(ii) characterisation of the molecular level interactions that occur between AFPs and AFGPs with lipids and thus provide insight into which compounds will protect and which will damage a particular membrane during chilling or freezing, and assist in the design of synthetic compounds tailored for interaction with specific membranes.

DNA Nanoshuttles: A New Class of DNA-Binding Agents

(with Dr L Rendina, University of Sydney)

The design of synthetic molecules that can interact with specific sequences and recognize local DNA conformations remains a high profile and exciting research area due to the enormous impact and diverse applications of these compounds in molecular biology, medicine and biotechnology. DNA Nanoshuttles are a new class of synthetic compounds containing platinum that mimic natural ring-shaped proteins and contain nanoscale dimensions that facilitate encircling of DNA by a threading mechanism (see figure on right). A study of the properties of DNA nanoshuttles will allow the potential of these compounds as a new class of DNA-binding agents to be evaluated and will expand the strategies available to chemists in designing new DNA-binding agents.

Medicinal Chemistry

(i) The antitumor metallocenes are a new class of organometallic anticancer agents, whose antitumour activity is believed to be related to interaction with DNA. In contrast to titanocene dichloride, the structurally related metallocenes (Cp₂MCl₂ M = V, Mo, Nb, Re) have been much less studied. The vastly different chemical stabilities of these complexes at physiological pH and different coordination chemistries of each complex point to significantly different mechanisms of antitumour action for each drug. Our recent studies have focused on establishing the mechanism of action of the less-well studied metallocenes. Systematic studies of the interaction(s) with cellular components including human serum albumin, glutathione, amino acids and blood plasma have been performed as well as micro-SRIXE (Synchroton Radiation Induced X-Ray Emission) to visualize the intracellular locations of the metals upon treatment of mammalian cells. These studies have allowed second generation derivatives of molybdocene dichloride predicted to have enhanced cellular uptake to be designed. The synthesis, biocoordination chemistry and antitumour activity studies of these new metallocene are currently in progress.

(ii) Streptonigrin is an aminoquinone antitumour antibiotic with broad spectrum activity against a range of cancers. The mechanism of antitumour action of the drug is believed to result from free radical-mediated DNA strand cleavage due to reductive activation of streptonigrin, in a process that involves metal ions and oxygen. Our research has focused on full characterisation of the structures, stabilities and redox chemistry of streptonigrin metal complexes of DNA and characterisation of the structures of streptonigrin-metal-DNA complexes allowing second generation analogues to be designed that will target DNA and topoisomerase enzymes in a different manner to the parent drug.

(iii) Saccharides have recently been identified as important DNA recognition motifs. We have examined the groove-binding characteristics of the saccharide side-chains present in the clinically important anthracyclines. Chemical and enzymatic footprinting has been complemented by NMR spectroscopic studies on the naturally occurring anthracyclines daunomycin, aclacinomycin and ditrisarubicin as well as a number of synthetic analogues. DNA-threaders that incorporate both major and minor groove binding sugars attached to an intercalating core, have also been designed and synthesised. The long DNA residence time of the these threaders is expected to modulate both DNA and topoisomerase interactions and hence biological activity.

(iv) DNA-Intercalators have been used to target the interaction of peptides and steroids with DNA. Intercalators have been attached to major groove binding alpha-helical peptides. A number of intercalator amino acids have been prepared and incorporated into the synthesis of DNA-bisintercalators of the general structure XSPSTSPZ, which are related to the heptad repeat present in RNA polymerase II. In addition, intercalator derivatives of cholic acid have been prepared with significant activity against L1210 mouse leukaemia cells observed with one derivative.

Metallosupramolecular Chemistry

The efficient assembly of chiral metallomacrocycles, capable of recognition of aromatic substrates, has been achieved. The 2,2'-bipyridyl coordination sites are important features that provide a mechanism whereby the dimensions of the cavities of the metallomacrocyles may be altered to accommodate optimal binding of aromatic substrates. Furthermore the reversible coordination chemistry and access to a range of metallomacrocycles of different shapes and dimensions (due to the range of metal geometries available) allows entry into virtual combinatorial libraries in which a substrate effectively templates or selects its optimal metallomacrocycle host from the range of geometries and oligomers present in solutions of our designed bisbipyridyl ligands and transition metal ions. Extension of these principles to allow the generation of diverse virtual combinatorial libraries encoded with the recognition features to potentially allow effective templating by a given guest molecule are currently under investigation.

Publications