Thinking Outside the Lattice
Though introductory textbooks often portray enzymes as static catalysts, in reality, protein dynamics are frequently just as important as structure. In the Ando lab, we study the domain motions, rearrangements, and quaternary structure transformations that proteins undergo during catalysis and regulation using a combination of biochemistry and biophysical tools. We focus on complex protein systems where we can apply our experience in technique development to propose novel solutions. We are especially interested in studying catalysis and regulation in metalloenzymes, proteins that use metal-containing cofactors to perform challenging chemistry.
As a structural enzymology lab, we are unique in that we often start with X-ray scattering, a structural technique that allows us to probe conformational disorder. For example, we use small-angle X-ray scattering (SAXS) to map the conformational landscapes of dynamic enzymes. By doing so, we not only gain intimate knowledge of the protein’s behavior in solution but also gain insight into experimental conditions that allow for hypothesis-driven structure determination by X-ray crystallography and cryo-electron microscopy (EM). Finally, we are challenging the notion that crystallography can only provide static snapshots. We are one of the few labs in the world harnessing information on correlated displacements contained in the diffuse scattering from protein crystals.
Diversity and Evolution of Complex Enzyme Allostery
Many important enzymes undergo complex regulation, often in the form of structural rearrangements. We are interested in understanding these allosteric mechanisms, the domains responsible, and the evolutionary pathways that have led to them. One such protein family of particular interest to us are the ribonucleotide reductases (RNRs), which are responsible in nearly all life for converting the building blocks of RNA to those of DNA. This enzyme family carries an ancient, structurally conserved catalytic core for performing radical chemistry but has diverged greatly in both radical generation and overall regulation. By applying a variety of structural techniques, we are interested in shedding light on the incredible diversity of allostery involved in this protein family.
Choreography of Highly Flexible Enzymes
Many enzymes of pharmaceutical interest are highly flexible or undergo significant domain rearrangements during catalysis. However, these systems often are conformationally heterogeneous and thus challenging to study with traditional crystallography. We use solution-scattering techniques in conjunction with other biophysical information to investigate the different conformations of these systems and the domain motions involved in catalysis. Ultimately, we hope to build dynamic models of these systems to better understand their function and regulation.
Correlated Motions in Protein Crystals
Conventional crystallography involves analyzing sharp diffraction patterns, commonly called Bragg data. However, real crystals are not perfectly periodic and produce additional scattering between the Bragg peaks. This smooth background pattern, known as diffuse scattering, contains information about correlated displacements within the crystal. With recent advances in detector technology, this weak signal can now be measured simultaneously with Bragg data. We are developing robust methods for collecting, processing, and analyzing diffuse scattering data from protein crystals that we ultimately will use to learn about correlated protein motions that are invisible to traditional crystallography.