Distinct functional surface regions on ubiquitin

The characterized functions of the highly conserved polypeptide ubiquitin a re to target proteins for proteasome degradation or endocytosis. The format ion of a polyubiquitin chain of at least four units is required for efficie nt proteasome binding. By contrast, monoubiquitin serves as a signal for th e endocytosis of plasma membrane proteins. We have defined surface residues that are important for ubiquitin's vital functions in Saccharomyces cerevi siae. Surprisingly, alanine scanning mutagenesis showed that only 16 of ubi quitin's 63 surface residues are essential for vegetative growth in yeast. Most of the essential residues localize to two hydrophobic clusters that pa rticipate in proteasome recognition and/or endocytosis. The others reside i n or near the tail region, which is important for conjugation and deubiquit ination. We also demonstrate that the essential residues comprise two disti nct functional surfaces: residues surrounding Phe(4) are required for endoc ytosis, whereas residues surrounding Ile(14) are required for both endocyto sis and proteasome degradation.