Structural Enzymology. Enzymes play a key role in all metabolic and cell-signaling processes. Characterization of an enzyme’s biological function must include the description of its mechanisms at an atomic level. Our laboratory is deciphering the catalytic mechanism of several enzyme families, using a combination of molecular biology, biochemistry and structural Biology. Systems under study fall into two classes: 1) Enzymes that recognize or process phosphates and 2) redox enzymes. These systems include: ATP-synthase, pyrophosphate hydrolases, farnesyl pyrophosphate synthases, PI3K, flavoenzymes, copper hydroxylases, and non-heme iron oxygenases. All experiments necessary to address mechanistic questions are carried out in the laboratory. Cloning and expression, ultrapurification, kinetic characterization, mutational analysis, mass spectrometry, crystallization, and structure determination by x-ray diffraction are some of the techniques we bring to bear to characterize the mechanisms of these enzymes. In addition to being intrinsically interesting some of these systems are being developed as targets for drug design.

Structural Thermodynamics. Most biological processes rely upon recognition and binding among macromolecules. We have developed several systems, such as anti-peptide antibodies and lectins that we are using to study protein-ligand interactions. As part of this research, we are developing computational methods to calculate the changes in the thermodynamic variables (ÄG, ÄH, ÄS) that take place when a protein recognized another macromolecule or a small ligand. Techniques used in this work involve monoclonal antibody development, x-ray diffraction and calorimetry, followed by empirical parameterization, and molecular mechanics/dynamics and statistical mechanics calculations. Results of these studies have a major impact on our understanding of binding energetics, including the estimation of binding affinities for structure-based drug design.

Publications:

Gabelli SB, Azurmendi HF, Bianchet MA, Amzel LM, Mildvan AS. (2006) X-ray, NMR, and mutational studies of the catalytic cycle of the GDP-Mannose mannosyl hydrolase reaction. Biochemistry 45(38):11290-303.

Gabelli, SB, McLellan, JS, Montalvetti, A, Oldfield, E, Docampo R. and Amzel, LM. (2005) Structure and mechanism of the farnesyl pyrophosphate synthase from Trypanosoma cruzi: implications for drug design. Proteins 62(1): 80-88.

L.M Amzel, Siebert X., Armstrong, A., Pabon, G. (2005) Thermodynamic calculations in biological systems. Biophys. Chem. 117:239-54.

Nadella M, Bianchet MA, Gabelli, SB, Barrila, J and Amzel, LM. (2005) Structure and activity of the axon guidance protein MICAL. Proc. Natl. Acad. Sci. 102: 16830-35.

Prigge ST, Eipper BA, Mains RE, Amzel LM (2004) Dioxygen binds end-on to mononuclear copper in a precatalytic enzyme complex. Science, 304: 864-867.