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Open Access Dissertation
Doctor of Philosophy (PhD)
Molecular and Cellular Biology
Year Degree Awarded
Month Degree Awarded
Dr. Daniel N. Hebert
Biochemistry | Cell Biology | Molecular Biology | Other Biochemistry, Biophysics, and Structural Biology | Other Cell and Developmental Biology
The ER Degradation-Enhancing Mannosidase-Like protein 1 (EDEM1) is a critical endoplasmic reticulum (ER) quality control factor involved in identifying and directing non-native proteins to the ER-associated protein degradation (ERAD) pathway. However, its recognition and binding properties have remained enigmatic since its discovery. Here we provide evidence for an additional redox-sensitive interaction between EDEM1 and Z/NHK that requires the presence of the single Cys on the α-1 antitrypsin ERAD clients. Moreover, this Cys-dependent interaction is necessary when the proteins are isolated under stringent detergent conditions, ones in which only strong covalent interactions can be sustained. This interaction is inherent to the mannosidase-like domain (MLD) of EDEM1, for which we demonstrate substrate-binding and glycan-trimming properties. EDEM1 appears to possess bipartite binding properties as it also binds these ERAD variants in a Cys-independent manner under milder detergent conditions. In addition, a second intrinsically disordered region (IDR) was identified and both are required for machinery interaction, but they are not necessary for Z/NHK binding. Interestingly, although the N-terminal IDR is required for binding to another ERAD substrate, in this case, neither IDR is required for Z/NHK binding, as the MLD, which lacks both IDRs, exhibits glycan-independent and redox-sensitive binding to Z/NHK. Not only is the MLD sufficient to preferentially interact with the α-1 antitrypsin ERAD clients, it also possesses, as demonstrated for the first time, catalytic properties as observed through glycan-trimming. Although the IDRs do not contribute to Z/NHK binding, they are required for binding to the oxidoreductase, ERdj5. These results expand our understanding of the range of binding interactions implemented by EDEM1 and help elucidate its role as an active mannosidase involved in the quality control machinery of the Endoplasmic Reticulum (ERQC).
Lamriben, Lydia, "IDENTIFYING FUNCTIONAL COMPONENTS OF THE ENDOPLASMIC RETICULUM QUALITY CONTROL AND DEGRADATION FACTOR EDEM1" (2018). Doctoral Dissertations. 1445.