Heme-oxidised IRP2 ubiquitin ligased-1 (HOIL-1), a constitutional part of the linear ubiquitin chain assembly complex (LUBAC), plays key roles in inflammatory and immune signalling. HOIL-1 deficient patients suffer from autoinflammation, immunodeficiency and amylopectinosis characterised by pathogenic accumulation of polyglucosan bodies in the patients’ muscles. HOIL-1 is a RING-between-RING (RBR) E3 ligase, catalysing ubiquitination in a distinct 2-step fashion where it first accepts ubiquitin from the E2 enzyme via transthiolation, then conjugates ubiquitin to the substrate. In stark contradiction to the notion that ubiquitin is linked to substrates on their Lys residues, HOIL-1 has been found to ubiquitinate non-canonical substrates, namely protein Ser/Thr residues and glucosaccharides, such as glycogen. However, deeper understanding of how HOIL-1 carries out these functions and if these functions are linked to the clinical phenotypes is hindered by lack of information on HOIL-1’s catalytic mechanism.
Here I present the crystal structure of HOIL-1 RBR caught in transthiolation with an E2-ubiquitin conjugate and an allosteric ubiquitin molecule. With structural and biochemical analysis, I further demonstrate how specific polyubiquitin chains allosterically regulate the transthiolation step. The structure reveals HOIL-1 C-terminal features that are distinct from the canonical RING2 fold and harbour a unique catalytic triad that may represent an adaption to HOIL-1’s atypical catalytic function. I will also present preliminary structural evidence which hints on how HOIL-1 RING2 may conjugate ubiquitin on maltose.
The current study promises future investigation on HOIL-1’s unique catalytic activity and substrate specificity. It also provides fundamental insights on deciphering HOIL-1’s function in physiological and pathological contexts.