Year of Publication

2017

Degree Name

Doctor of Philosophy (PhD)

Document Type

Doctoral Dissertation

College

Arts and Sciences

Department

Chemistry

First Advisor

Dr. Anne-Frances Miller

Abstract

Gln146 is a highly conserved outer-sphere amino acid residue at the active site of MnSOD. It serves as a hydrogen bond donor to both the solvent molecules at the active site and Tyr 34, the conserved “gateway” amino acid residue. This dissertation develops our understanding of the effect of amino acid Gln146 at the second shell of the active site of metalloprotein MnSOD in facilitating metal binding, the modulation of redox potential, and the optimization of catalytic activity and structure stability. Different from the wild-type MnSOD, Q146E is always purified as a completely apo-protein with zero active metal ion and non-catalytic activity. But unlike apoMn-SOD, Q146E as an apo-protein exhibits extraordinarily conformational restricted protein structure. Because of the hyper-thermal stability of Q146E-apoMn-SOD, the protein itself loses its ability to form an “open” state which is responsible for the metal binding and protein maturation of apoMn-SOD. Increased thermal stability of protein structure also can be found in other mutants at the same position Gln146, Q146X-MnSOD (X=A, C, N or S), in addition to decreased catalytic activity, low Mn3+/Mn ratio and the reduction potential of the activity site. Thus Gln146 is very effective in facilitating metal binding by destabilizing protein structure.

Digital Object Identifier (DOI)

https://doi.org/10.13023/ETD.2017.049

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