Different HECT domain ubiquitin ligases employ distinct mechanisms of polyubiquitin chain synthesis.
Individual ubiquitin (Ub)-protein ligases (E3s) cooperate with specific Ub-conjugating enzymes (E2s) to modify cognate substrates with polyubiquitin chains. E3s belonging to the Really Interesting New Gene (RING) and Homologous to E6-Associated Protein (E6AP) C-Terminus (HECT) domain families utilize distinct molecular mechanisms. In particular, HECT E3s, but not RING E3s, form a thiol ester with Ub before transferring Ub to the substrate lysine. Here we report that different HECT domain E3s can employ distinct mechanisms of polyubiquitin chain synthesis. We show that E6AP builds up a K48-linked chain on its HECT cysteine residue, while KIAA10 builds up K48- and K29-linked chains as free entities. A small region near the N-terminus of the conserved HECT domain helps to bring about this functional distinction. Thus, a given HECT domain can specify both the linkage of a polyubiquitin chain and the mechanism of its assembly.
Wang
Min
M
Department of Biochemistry and Molecular Biology, Bloomberg School of Public Health, Johns Hopkins University, Baltimore, MD 21205, USA.
Pickart
Cecile M
CM
D001665:Binding Sites; D015153:Blotting, Western; D003001:Cloning, Molecular; D003545:Cysteine / Q000737:chemistry; D018076:DNA, Complementary / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D008239:Lysine / Q000737:chemistry; D009097:Multienzyme Complexes / Q000737:chemistry; D010957:Plasmids / Q000378:metabolism; D025821:Polyubiquitin / Q000737:chemistry*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D013997:Time Factors; D025801:Ubiquitin / Q000737:chemistry; D044763:Ubiquitin-Conjugating Enzymes / Q000737:chemistry; D043743:Ubiquitin-Protein Ligase Complexes / Q000737:chemistry; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry*
D018076:DNA, Complementary; D009097:Multienzyme Complexes; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D025801:Ubiquitin; D025821:Polyubiquitin; D044763:Ubiquitin-Conjugating Enzymes; D043743:Ubiquitin-Protein Ligase Complexes; C471639:UBE3A protein, human; D044767:Ubiquitin-Protein Ligases; D005982:Glutathione Transferase; D008239:Lysine; D003545:Cysteine