Lab Members

 

 Research

My laboratory explores specific aspects of microbial physiology and enzymology related to transition metals. In particular, we study mechanisms of catalysis by metalloenzymes and characterize the biosynthesis of protein metallocenters. We use an array of experimental techniques and approaches that ranges from gene cloning to enzyme kinetics, from site-directed mutagenesis to metal ion binding assays, and from active site peptide studies to biophysical spectroscopic methods.

A major emphasis in my laboratory focuses on characterization of several ferrous ion and alpha-ketoglutarate dependent hydroxylases. TauD functions in bacterial metabolism of sulfonated compounds and has become the paradigm of this enzyme family because of our spectroscopic, mutagenic and crystallographic studies. XanA is a fungal enzyme that metabolizes xanthine by a novel mechanism . A trypanosomal enzyme is required for the synthesis of base J in these protozoa. Finally, several projects involve unique DNA-repair enzymes. Current work with these fascinating enzymes includes metallocenter analysis by spectroscopic methods, characterization of site-directed mutant proteins and examination of alternate substrates and inhibitors.

A second area of emphasis in the laboratory centers on the mechanism of activation for the nickel-dependent enzyme urease. Bacterial urease is associated with the formation of urinary deposits (kidney stones) during human infection, and uncontrolled hydrolysis of urea based fertilizers can lead to crop damage; thus, a detailed understanding of urease activation may allow the design of pharmacologically or agriculturally effective inhibitors of this enzyme. Within the urease gene cluster are seven genes: three genes encode the three urease subunits (of known 3D structure), and four genes encode "accessory proteins". One of the accessory proteins is a "metallochaperone" that delivers nickel ion; we now know the structure of this protein. The other three accessory proteins work together to form a complex with urease apoprotein. We are presently examining the role of each of these components in the mechanism of nickel incorporation into urease.