With this award from the Major Research Instrumentation (MRI) Program that is cofunded by Chemistry, Professor Karen Allen and colleagues from Boston University will acquire a circular dichroism (CD) spectrometer. The proposal is aimed at enhancing research training and education at all levels, especially in areas such as (a) chemical and catalytic mechanisms of enzymes, (b) characterization of proteins, (c) natural product biosynthesis, (d) assignment of absolute stereochemistry of enantiomerically pure compounds, (e) determination of structural perturbations caused by mutations in glycolytic enzyme, and (f) stabilization of the functional form of NF-kappa-b essential modulator (NEMO) protein.
A circular dichroism spectrophotometer will produce spectral data that provide information on molecular structure and especially the conformations of bio-molecules such as nucleic acids, proteins, and enzymes. The principle of measurement is based on differential absorption of left and right circularly polarized light by optically active samples such as those in the double helix of DNA. The instrument will provide research and laboratory training in analytical techniques that allow measurement of physical properties of organic and inorganic molecules to graduate and undergraduate students as well as postdoctoral fellows. The CD spectrophotometer will be integrated into training of students in the Chemistry and Biology Departments, as well as the Biochemistry and Molecular Biology Program for the study of intermolecular interactions, geometric isomers and protein structure.
Intellectual Merit This award supported the purchase of a new circular dichroism (CD) Applied Photophysics CS/2 Chirascan spectrometer in the Chemical Instrumentation Center at Boston University (BU). The goal of the acquisition of the CD instrument is to support and enhance National Science Foundation and other agency-funded activities at BU. Often, these activities require the production of proteins outside their native environment at scales required for biophysical and biochemical measurements. The research also sometimes necessitates the production of chemically modified proteins. The use of CD in these systems allows measurements to ensure that the protein retains the same structure as when it is produced its native state and environment. Additional features of the instrument include a titration apparatus that enables the quantitative assessment of the stability of target proteins and allows the assessment of differences in stability between modified and unmodified forms of the protein. The stop-flow capability enables the same measurements to be achieved at different time points, allowing the determination of changes in protein structure over time (protein dynamics). The coupling of dynamics with binding events has recently become a central emphasis in the areas of drug discovery, signaling processes in the cell, regulation of enzymes, and enzyme-mediated catalysis. CD instrument use at BU has enhanced research resulting in six primary research publications. Highlights of the research enabled by the instrument include 1) determination of the structure of an enzyme key to the organism that is the causative agent of elephantitis 2) generation of an experimentally tractable and native-like form of the protein NEMO that plays important roles in cancer, immunity, and inflammation and 3) the assessment of the structure and the link to function in the protein MitoNEET that is suggested to play roles in the metabolism of iron and in coordinating energy production and utilization in the cell. Broader Impact As envisioned, the use of the instrument has also served in education activities. The CD has been utilized at BU in the course CH303 Instrumental Analysis since the course’s inception in Fall 2013. Using a series of standard solutions of known concentration, the students use CD measurements to determine the purity of an unknown compound. Additionally, the use of the CD instrument in research activities has provided training for both graduate (Ph.D. level), undergraduate, and high-school students. Specifically, lead graduate students have been trained who in turn have trained undergraduates and high-school students in instrument use. These include students from the summer Research Internship in Science & Engineering Program (RISE) program that provides an opportunity for academically motivated rising high school seniors to conduct university-level research in state-of-the-art laboratories. Undergraduate students include those from the Summer Research Experiences for Undergraduates (REU) program where research is conducted by outstanding students from other colleges and universities, particularly those without plentiful research opportunities such as community colleges. REU is designed to encourage graduate education in the sciences for talented undergraduate students, especially those from populations traditionally underrepresented in the sciences. Lastly, the instrument has been used in a workshop, organized in collaboration with Applied Photophysics, the instrument vendors, to teach the theory and introduce the uses of CD in the study of proteins. The workshop was attended by BU users as well as scientists and students from companies and other universities and colleges in Massachusetts, Connecticut and New York, allowing broad dissemination of knowledge.
