Studying the Structure and Interactions of the Globin-Coupled Sensor in P. carotovorum
Restricted (Penn State Only)
- Author:
- Potter, Jacob
- Graduate Program:
- Biochemistry, Microbiology, and Molecular Biology
- Degree:
- Doctor of Philosophy
- Document Type:
- Dissertation
- Date of Defense:
- December 14, 2022
- Committee Members:
- Lorraine Santy, Major Field Member
Scott Showalter, Outside Unit & Field Member
Timothy Miyashiro, Major Field Member
Timothy Meredith, Major Field Member
Wendy Hanna-Rose, Program Head/Chair
Emily Weinert, Chair & Dissertation Advisor - Keywords:
- Biochemistry
Microbiology
Molecular Biology
SAXS
Pectobacterium
PccGCS
Diguanylate cyclase
c-di-GMP
cryo-EM
globin
globin coupled sensor
oxygen sensing
structural biology
enzyme
oligomer
domain
signal transduction
bacteria
virulence
cross linking
spectroscopy - Abstract:
- Bacteria frequently utilize globin-coupled sensors (GCS) to sense and react to diatomic gases like O2¬ and CO. These GCS include a heme-containing globin coupled to one of many potential output domains that allow for expression of various phenotypes. Of particular interest is the diguanylate cyclase (DGC) coupled GCS from Pectobacterium carotovorum spp. carotovorum WPP14 (termed PccGCS), a plant pathogen. DGC-containing GCS synthesize the bacterial second-messenger, cyclic dimeric guanosine monophosphate (c-di-GMP) and are implicated in a variety of cellular phenotypes such as motility, biofilm formation, and even virulence. In this dissertation, I investigate the structure and interactions of PccGCS while utilizing a variety of techniques, including small-angle X-ray scattering (SAXS), cryo-EM, enzymatic activity assays, cross-linking, and spectroscopy. Examination within these chapters has revealed much on relationship of PccGCS DGC activity with oligomeric state and domain mobility. From this, several important residues have been determined that shed light into the mechanism of signal transduction across the domains of this molecule. Additionally, work from multiple chapters in this work contain data that will set the stage for the pursuit of important protein-protein interactions, as well as the elusive first high-resolution structure of a GCS protein. Finally, further research into globin-coupled sensors will expand our knowledge of important interactions involved in bacterial O2, which may lead to therapeutics that modulate virulence in related human and agricultural pathogens.