Distinct functional surface regions on ubiquitin.
The characterized functions of the highly conserved polypeptide ubiquitin are to target proteins for proteasome degradation or endocytosis. The formation of a polyubiquitin chain of at least four units is required for efficient proteasome binding. By contrast, monoubiquitin serves as a signal for the endocytosis of plasma membrane proteins. We have defined surface residues that are important for ubiquitin's vital functions in Saccharomyces cerevisiae. Surprisingly, alanine scanning mutagenesis showed that only 16 of ubiquitin's 63 surface residues are essential for vegetative growth in yeast. Most of the essential residues localize to two hydrophobic clusters that participate in proteasome recognition and/or endocytosis. The others reside in or near the tail region, which is important for conjugation and deubiquitination. We also demonstrate that the essential residues comprise two distinct functional surfaces: residues surrounding Phe(4) are required for endocytosis, whereas residues surrounding Ile(44) are required for both endocytosis and proteasome degradation.
Sloper-Mould
K E
KE
Department of Biochemistry, Molecular Biology and Cell Biology, Northwestern University, Evanston, Illinois 60208, USA.
Jemc
J C
JC
Pickart
C M
CM
Hicke
L
L
D000409:Alanine / Q000737:chemistry; D000595:Amino Acid Sequence; D001665:Binding Sites; D002455:Cell Division; D003546:Cysteine Endopeptidases / Q000378:metabolism; D004705:Endocytosis; D007532:Isoleucine / Q000737:chemistry; D000072235:Mating Factor; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010455:Peptides / Q000378:metabolism; D010649:Phenylalanine / Q000737:chemistry; D010957:Plasmids / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex; D011485:Protein Binding; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D013997:Time Factors; D014452:Ubiquitins / Q000737:chemistry* / Q000502:physiology*; D014867:Water / Q000378:metabolism
D009097:Multienzyme Complexes; D010455:Peptides; D011993:Recombinant Fusion Proteins; D014452:Ubiquitins; D007532:Isoleucine; D014867:Water; D010649:Phenylalanine; D000072235:Mating Factor; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; D000409:Alanine