Bharati Mitra, Ph.D..

We are interested in the catalytic mechanisms employed by enzymes, with emphasis on the active site structures responsible for the rate accelerations and substrate specificity achieved by enzymes. We have two ongoing projects. In one, we are studying metal ion homeostasis of essential trace metals as well resistance mechanisms to toxic metals in the cell, with special emphasis on zinc. Specifically, we are interested in the role played by transporters located in membranes in maintaining a balanced concentration of essential metals including zinc, cobalt, copper and nickel, as well as in mediating resistance to metals such as lead, cadmium and mercury. In particular, we are investigating the relationship between structure and function for a class of transporters that couple ATP hydrolysis and soft-metal translocation across membranes. Another long-standing project in our laboratory involves the reaction mechanism of a large family of FMN-containing proteins. These enzymes, present in bacteria, archea and eukarya, including humans, oxidize 2-hydroxyacids to2-ketoacids using a variety of oxidants such as oxygen, heme and quinones, depending on their location inside the cell. We are investigating the overall reaction mechanism, as well as the basis of substrate specificity and most importantly, the control of reactivity to molecular oxygen in this flavoprotein family. Techniques used in our laboratory include genetic engineering, membrane protein characterization, steady-state and stopped-flow kinetics, absorbance and fluorescence spectroscopy, binding studies, and protein structure determination.

Dewanti, A.R., Mitra, B. A Transient Intermediate in the Reaction Catalyzed by (S)-Mandelate Dehydrogenase. In press, Biochemistry 42, 2003.

Hou, Z., Mitra, B. Characterization of the metal specificity of ZntA from Escherichia coli using the acylphosphate intermediate. J. Biol. Chem. 278:28455-28461, 2003.

Xu, Y., Dewanti A.R., Mitra, B. The Arginine165/Arginine277 pair in (S)-Mandelate Dehydrogenase from Pseudomonas putida: role in catalysis and substrate binding. Biochemistry 41:12313-12319, 2002.

Hou, Z-j., Narindrasorasak, S., Bhushan, B., Sarkar B., Mitra B. Functional analysis of chimeric proteins of the Wilson Cu(I)-transporting ATPase (ATP7B) and ZntA, a Pb(II)/Zn(II)/Cd(II)-translocating ATPase from Escherichia coli. J. Biol. Chem. 276:40858-40863, 2001.

Sukumar, N., Xu, Y., Gatti, D.L., Mitra B., Mathews, F.S. Structure of an Active, Soluble Mutant of the Membrane-Associated (S)-Mandelate Dehydrogenase. Biochemistry 40: 9870-9878, 2001.

Mitra, B., Sharma, R. The cysteine-rich amino-terminal domain of ZntA, a Pb(II)/Cd(II)/Zn(II)-translocating ATPase from Escherichia coli, is not essential for its function. Biochemistry 40:7694-7699, 2001.

Lehoux, I.E., Mitra, B. The Role of Arginine277 in (S)-Mandelate Dehydrogenase from Pseudomonas putida in Substrate Binding and Transition State Stabilization. Biochemistry 39:10055-10065, 2000.