Theses and Dissertations


He Song

Issuing Body

Mississippi State University


Emerson, Joseph P.

Committee Member

Lewis, Edwin A.

Committee Member

Mlsna, Todd E.

Committee Member

Henry, William P.

Committee Member

Zhang, Dongmao

Date of Degree


Document Type

Dissertation - Open Access



Degree Name

Doctor of Philosophy


College of Arts and Sciences


Department of Chemistry


Carbonic anhydrase (CA) is a well-studied, zinc dependent metalloenzyme that catalyzes hydrolysis of carbon dioxide to the bicarbonate ion. In the past, metal binding studies related to CA have continually relied on equilibrium dialysis measurements to ascertain an extremely strong association constant (Ka= approx. 1.2 x 1012) for Zn2+. However, new methodology has allowed us to collect data using isothermal titration calorimetry (ITC), which calls that number and the association constants for many other first row transition metal ions into question. Thermodynamic parameters associated with Zn2+, Cu2+, Ni2+, and Co2+ binding to apoCA are unraveled from a series of complex equilibria associated with the in vitro metal binding event. This in-depth analysis adds clarity to the complex ion chemistry associated with metal ion binding to carbonic anhydrase and validates thermochemical methods that accurately measure association constants and thermodynamic parameters for complex-ion and coordination chemistry observed in vitro. Additionally, the as-isolated and the reconstituted ZnCA and other metalsubstituted CAs were probed using X-ray absorption spectroscopy. Both X-ray absorption near edge structure (XANES) and extended X-ray absorption fine structure (EXAFS) analyses indicate the metal centers in the reconstituted carbonic anhydrases offer new metal binding coordination sites that can be used as models to understand nonheme metal sites in vivo.