The Dynamics of Circulatory Zinc Handling and Transport

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There are 25 chemical elements that are required for mammalian life, 15 of these elements are metals. Zinc, in its ionised form Zn2+, is an essential metal ion in mammals and performs a wide range of important physiological functions by allowing many vital chemical reactions to occur. Zinc is known to play roles in fertility and development, the immune system, ageing, and major diseases such as Alzheimer's, diabetes, and cancer. Unfortunately however, the molecular mechanisms that enable zinc to get to where it is needed after it enters the bloodstream, following sequestration from the diet, are not fully understood.It is known that a particular protein called serum albumin (that is highly abundant in the blood) plays an important role in transporting zinc (and other metals including calcium and magnesium) in the blood. Serum albumin not only carries zinc through the circulatory system but also mediates its uptake into cells. The molecular mechanisms that control the interaction between zinc and serum albumin and its uptake into cells are largely unknown. It is important that these interactions are understood. Albumin also transports other types of molecules (e.g. fatty acids, hormones) and binding of one particular molecule can affect binding of another at a separate site. For example, we previously identified a zinc site that is perturbed by fatty acid-binding elsewhere on the molecule. Physiological events that alter the small molecule composition of blood can therefore alter metal transport processes. Such events may be short-term (e.g. fasting, eating, infection, stroke) or long-term (obesity, disease). Long-term alterations in blood chemistry are particularly likely to have serious consequences due to the knock-on effects caused by altered metal binding/delivery.We have identified three regions on the molecule that are likely to form novel metal binding sites. We will examine whether these regions that are involved in or impact upon zinc transport by synthesising mutant serum albumin proteins with chemical alterations at each site. These alterations will remove important chemical groups that are likely to participate in metal binding. The ability of each "altered protein" to bind zinc and other relevant metals will be examined and compared to the native (or "normal") protein using complementary approaches. We will also examine whether binding of molecules (including metals) at other sites alters the binding properties of each site.The importance of known and newly identified zinc-binding sites in mediating zinc uptake into cultured vascular endothelial cells (a particular type of cell that lines the blood vessels) will be determined by incubating these cells with albumins that contain alterations at each zinc site and physiological levels of zinc. The amount of zinc taken up by the cells will then be measured. The outcome of these experiments will allow a better understanding of how metal ions are transported in the circulatory system. Particularly with regard to the events that can alter this process and the importance of albumin in mediating cellular zinc uptake. This provides important physiological information that will help us to understand a variety of physiological and disease processes that involve metals and will aid in their study.