THE UBIQUITIN-LIKE PROTEIN SMT3P IS ACTIVATED FOR CONJUGATION TO OTHER PROTEINS BY AN AOS1P UBA2P HETERODIMER/

SMT3 is an essential Saccharomyces cerevisiae gene encoding a 11.5 kDa protein similar to the mammalian ubiquitin-like protein SUMO-1. We ha ve found that Smt3p, like SUMO-1 and ubiquitin, can be attached to oth er proteins post-translationally and have characterized the processes leading to the activation of the Smt3p C-terminus for conjugation. Fir st, the SMT3 translation product is cleaved endoproteolytically to exp ose Gly98, the mature C-terminus, The presence of Gly98 is critical fo r Smt3p's abilities to be conjugated to protein substrates and to comp lement the lethality of a smt3 Delta strain, Smt3p undergoes ATP-depen dent activation by a novel heterodimeric enzyme consisting of Uba2p, a previously identified 71 kDa protein similar to the C-terminus of ubi quitin-activating enzymes (E1s), and Aos1p (activation of Smt3p), a 40 kDa protein similar to the N-terminus of E1s. Experiments with condit ional uba2 mutants showed that Uba2p is required for Smt3p conjugation in vivo, Furthermore, UBA2 and AOS1 are both essential genes, providi ng additional evidence that they act in a distinct pathway whose role in cell viability is to conjugate Smt3p to other proteins.