Molecular basis for lysine specificity in the yeast ubiquitin-conjugating enzyme Cdc34.
Ubiquitin (Ub)-conjugating enzymes (E2s) and ubiquitin ligases (E3s) catalyze the attachment of Ub to lysine residues in substrates and Ub during monoubiquitination and polyubiquitination. Lysine selection is important for the generation of diverse substrate-Ub structures, which provides versatility to this pathway in the targeting of proteins to different fates. The mechanisms of lysine selection remain poorly understood, with previous studies suggesting that the ubiquitination site(s) is selected by the E2/E3-mediated positioning of a lysine(s) toward the E2/E3 active site. By studying the polyubiquitination of Sic1 by the E2 protein Cdc34 and the RING E3 Skp1/Cul1/F-box (SCF) protein, we now demonstrate that in addition to E2/E3-mediated positioning, proximal amino acids surrounding the lysine residues in Sic1 and Ub are critical for ubiquitination. This mechanism is linked to key residues composing the catalytic core of Cdc34 and independent of SCF. Changes to these core residues altered the lysine preference of Cdc34 and specified whether this enzyme monoubiquitinated or polyubiquitinated Sic1. These new findings indicate that compatibility between amino acids surrounding acceptor lysine residues and key amino acids in the catalytic core of ubiquitin-conjugating enzymes is an important mechanism for lysine selection during ubiquitination.
Sadowski
Martin
M
Cell Cycle and Cancer Unit, St. Vincent's Institute of Medical Research, 9 Princes St., Fitzroy, Melbourne, Victoria 3065, Australia.
Suryadinata
Randy
R
Lai
Xianning
X
Heierhorst
Jörg
J
Sarcevic
Boris
B
D000595:Amino Acid Sequence; D064173:Anaphase-Promoting Complex-Cyclosome; D049109:Cell Proliferation; D002470:Cell Survival; D050756:Cyclin-Dependent Kinase Inhibitor Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008239:Lysine / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D044843:SKP Cullin F-Box Protein Ligases / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000502:physiology; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D013379:Substrate Specificity / Q000235:genetics; D025801:Ubiquitin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D044763:Ubiquitin-Conjugating Enzymes / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D043743:Ubiquitin-Protein Ligase Complexes / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination
D050756:Cyclin-Dependent Kinase Inhibitor Proteins; C086926:SIC1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D025801:Ubiquitin; C577449:CDC34 protein, S cerevisiae; C577447:CDC34 protein, human; D044763:Ubiquitin-Conjugating Enzymes; D043743:Ubiquitin-Protein Ligase Complexes; D064173:Anaphase-Promoting Complex-Cyclosome; D044843:SKP Cullin F-Box Protein Ligases; D008239:Lysine