TY  - GEN
N2  - Here, I present a comprehensive assay termed UbiReal for monitoring each step of the Ub signaling cascade, including linkage-specific polyUb ligation and deubiquitination. Next, I use bioinformatic tools and recombinant protein expression to identify additional, HECT-like ligases from human and plant pathogens, and validate these ligases using UbiReal. I apply structural biology and biochemical techniques to elucidate the mechanism of Ub ligation by the bacterial HECT-like ligases, and capture a rare glimpse of a linkage-specific polyUb ligation interface. I leverage insights from the linkage-specific polyUb interface to engineer an NleL specific for K6-linked polyUb, and show that similar rules influence the ligation of K6-linked polyUb in humans. Finally, through analysis of a family of deubiquitinating enzymes in bacteria, I demonstrate the utility of the deubiquitinase LotA as a tool for detecting the presence of K6-linked polyUb. Collectively, this dissertation provides the critical tools for understanding K6-linked polyUb biology, and in particular sets the stage for elucidating the enigmatic role of K6-linked polyUb at the host-pathogen interface.
DO  - 10.6083/bpxhc42360
DO  - doi
AB  - Here, I present a comprehensive assay termed UbiReal for monitoring each step of the Ub signaling cascade, including linkage-specific polyUb ligation and deubiquitination. Next, I use bioinformatic tools and recombinant protein expression to identify additional, HECT-like ligases from human and plant pathogens, and validate these ligases using UbiReal. I apply structural biology and biochemical techniques to elucidate the mechanism of Ub ligation by the bacterial HECT-like ligases, and capture a rare glimpse of a linkage-specific polyUb ligation interface. I leverage insights from the linkage-specific polyUb interface to engineer an NleL specific for K6-linked polyUb, and show that similar rules influence the ligation of K6-linked polyUb in humans. Finally, through analysis of a family of deubiquitinating enzymes in bacteria, I demonstrate the utility of the deubiquitinase LotA as a tool for detecting the presence of K6-linked polyUb. Collectively, this dissertation provides the critical tools for understanding K6-linked polyUb biology, and in particular sets the stage for elucidating the enigmatic role of K6-linked polyUb at the host-pathogen interface.
AD  - Oregon Health and Science University
T1  - Strangers in a strange land: bacteria provide mechanistic insight into eukaryotic ubiquitination
DA  - 2023-11-24
AU  - Franklin, Tyler
L1  - https://digitalcollections.ohsu.edu/record/42360/files/permission-LotA.pdf
L1  - https://digitalcollections.ohsu.edu/record/42360/files/permission-PPUbireal.pdf
L1  - https://digitalcollections.ohsu.edu/record/42360/files/Franklin.Tyler.2023.pdf
PB  - Oregon Health and Science University
LA  - eng
PY  - 2023-11-24
ID  - 42360
L4  - https://digitalcollections.ohsu.edu/record/42360/files/permission-LotA.pdf
L4  - https://digitalcollections.ohsu.edu/record/42360/files/permission-PPUbireal.pdf
L4  - https://digitalcollections.ohsu.edu/record/42360/files/Franklin.Tyler.2023.pdf
KW  - Biochemistry
KW  - Microbiology
KW  - Crystallography, X-Ray
KW  - ubiquitin
KW  - e3 ubiquitin ligase
KW  - deubiquitinase
KW  - bacteria
KW  - nlel
KW  - sopa
KW  - lota
TI  - Strangers in a strange land: bacteria provide mechanistic insight into eukaryotic ubiquitination
Y1  - 2023-11-24
L2  - https://digitalcollections.ohsu.edu/record/42360/files/permission-LotA.pdf
L2  - https://digitalcollections.ohsu.edu/record/42360/files/permission-PPUbireal.pdf
L2  - https://digitalcollections.ohsu.edu/record/42360/files/Franklin.Tyler.2023.pdf
LK  - https://digitalcollections.ohsu.edu/record/42360/files/permission-LotA.pdf
LK  - https://digitalcollections.ohsu.edu/record/42360/files/permission-PPUbireal.pdf
LK  - https://digitalcollections.ohsu.edu/record/42360/files/Franklin.Tyler.2023.pdf
UR  - https://digitalcollections.ohsu.edu/record/42360/files/permission-LotA.pdf
UR  - https://digitalcollections.ohsu.edu/record/42360/files/permission-PPUbireal.pdf
UR  - https://digitalcollections.ohsu.edu/record/42360/files/Franklin.Tyler.2023.pdf
ER  -