ORCID

http://orcid.org/0000-0003-2660-9466

Date of Award

Summer 8-15-2021

Author's School

Graduate School of Arts and Sciences

Author's Department

Biology & Biomedical Sciences (Computational & Molecular Biophysics)

Degree Name

Doctor of Philosophy (PhD)

Degree Type

Dissertation

Abstract

Proteins are macromolecular machines that play a role in nearly every biological process. They are dynamic molecules which adopt many different conformations as they fold into their 3D structures, interact with their binding partners, and perform their functions. The most probable (lowest energy) protein conformation is referred to as the ground state, and this is often assumed to be the state determined by experimental methods such as x-ray crystallography. However, proteins also adopt higher energy excited states which can have significant probabilities. As these excited states are notoriously difficult to find and study, it is unclear if excited states contribute to the protein’s function and ultimately to the organism’s survival. To investigate this question, I examine the excited states of β-lactamases, enzymes which confer bacteria with antibiotic resistance by degrading β-lactam antibiotics such as penicillin. I first consider a subset of excited states that contain a cryptic pocket, or a pocket which is absent in the ground state conformation. I find that the Ω-loop cryptic pocket seen in TEM β-lactamase plays a role in the protein’s ability to hydrolyze different classes of antibiotics and specifically that the population of open pocket conformations is predictive of activity against the substrate benzylpenicillin. I next consider the subset of excited states along the protein folding pathway. I find that β-lactamase stability contributes significantly to the ability of bacteria expressing β-lactamase to survive in the presence of antibiotic and specifically that including the population of folding intermediate conformations improves the predictive power of my model for bacterial fitness. These results demonstrate our understanding of how excited states are connected to β-lactamase function and survival of bacteria expressing β-lactamases. This work also helps establish the utility of cryptic pockets as drug targets and aids in our understanding of how mutations (genotype) result in changes in fitness (phenotype).

Language

English (en)

Chair and Committee

Gregory R. Bowman

Committee Members

Gautam Dantas, Michael Greenberg, Linda Pike, Christina Stallings,

Download

Share

COinS