UC San Diego

Endoplasmic Reticulum Associated Degradation (ERAD) is a conserved pathway that is responsible for the degradation of aberrant proteins in the ER. These proteins are first tagged with ubiquitin through the concerted action of E1, E2, and E3 ubiquitination enzymes and then degraded by the proteasome. Hrd1p is one of the principle E3 ubiquitin ligases involved in ERAD. It is an integral ER membrane protein with six transmembrane spans and a cytosolic RING domain and is responsible for the recognition and degradation of a wide variety of ER-localized misfolded proteins, including lumenal (ERAD-L) and integral membrane (ERAD-M) substrates. Previous studies have suggested that Hrd1p substrate recognition function as an "allosteric" model and that the Hrd1p transmembrane domain is a key site for substrate selectivity. To further explore this idea, we have conducted a screen to isolate Hrd1p mutants that show increased activity toward substrates of the HRD pathway. We have isolated a single point mutation in the transmembrane domain of Hrd1p that imparts more effective degradation of an ERAD-M substrate. Further analysis of this mutation revealed that it causes selective increased activity toward a single substrate and degrades all other substrates normally. This selective increase in activity supports the idea that different regions of the transmembrane domain are responsible for the recognition of different Hrd1p substrates