Spectroscopic Studies of Molybdenum and Tungsten Enzymes

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Transition metal ions are integral to a large number of enzymes and other proteins, with transition metalloproteins constituting about 11% of a typical organism’s genomic output. Transition metal ions are used by the enzymes to catalyze many of the most chemically challenging and industrially relevant reactions carried out by living organisms. Among all of the transition metals employed by living things, molybdenum and tungsten are distinguished as being the only second-row and third-row transition elements with a known function in life. Tungsten is further distinguished as the heaviest element known to have functional roles in biology. The structures of molybdenum and tungsten enzymes are very closely related, both containing an unusual sulfur-containing organic cofactor called molybdopterin. The roles of molybdenum and tungsten in metalloenzymes often appear to be interchangeable, with molybdenum enzymes found broadly in higher organisms, while the role of molybdenum is filled by tungsten in more primitive organisms. Together, the molybdenum and tungsten enzymes have important roles in almost all living organisms, from archaea to bacteria through plants to invertebrates and mammals. They do this within enzyme active sites that, while sharing common structural motifs such as the molybdopterin, are remarkable in the range of different chemical reactions are catalyzed. The functional roles fulfilled by the enzymes are equally diverse; for example, they play essential roles in microbial respiration, in the uptake of nitrogen in green plants, in controlling insect eye color, and in human health. The tungsten enzymes are thought to have played important roles in early life on earth, most notably in the organism known as the LUCA (last universal common ancestor), sometimes called the cenancestor, the progenitor of all earthly life now extant. The proposed work is a systematic study of the major families of molybdenum and tungsten enzymes using a combination of synchrotron-based and computational chemistry methods. We propose to further develop a novel holistic approach in combining information from X-ray absorption spectroscopy, advanced computer molecular modeling, and crystal structures to obtain the most accurate picture yet of the active sites, and thereby to elucidate their detailed catalytic mechanisms. The ultimate goal of the work is thus to develop an in-depth quantitative structural and electronic understanding of the catalytic mechanisms used by the enzymes, comparing and contrasting between and within the different families. This will lead to a richer understanding of how molybdenum and tungsten enzymes achieve their remarkable functional diversity. The benefits of this research are four-fold: (i) The results will yield detailed insights for an important group of enzymes. (ii) The holistic approach and the methods developed for it will be broadly applicable to studies of any metalloprotein. (iii) The results will provide insights into a group of enzymes that were important in primordial life on earth. (iv) By driving and developing techniques at the Canadian Light Source, the research will strengthen the opportunity for Canadian researchers to utilize these synchrotron tools in structural molecular biology.