Biochemical analysis of Angelman syndrome-associated mutations in the E3 ubiquitin ligase E6-associated protein.
Angelman syndrome is a severe neurological disorder characterized by mental retardation, absent speech, ataxia, seizures, and hyperactivity. The gene affected in this disorder is UBE3A, the gene encoding the E6-associated protein (E6AP) ubiquitin-protein ligase. Most patients have chromosomal deletions that remove the entire maternal allele of UBE3A. However, a small subset of patients have E6AP point mutations that result in single amino acid changes or short in-frame deletions that still allow translation of a full-length protein. By studying these point mutations in E6AP, we found a strong correlation between Angelman-associated mutations and a loss of E3 ubiquitin ligase activity. Interestingly the point mutations affect E6AP activity in different ways. Some mutant proteins cannot form thiol ester intermediates with ubiquitin, others retain the thiol ester formation activity but cannot efficiently transfer ubiquitin to a substrate, and still others are unstable in cells. Our results suggest that the loss of E6AP catalytic activity and likely the improper regulation of E6AP substrate(s) are important in the development of Angelman syndrome.
Cooper
Eric M
EM
Harvard Medical School, Department of Pathology, Boston, Massachusetts 02115, USA.
Hudson
Amy W
AW
Amos
Joseph
J
Wagstaff
Joseph
J
Howley
Peter M
PM
D017204:Angelman Syndrome / Q000235:genetics*; D000818:Animals; D001667:Binding, Competitive; D002384:Catalysis; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000378:metabolism; D004952:Esters / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism; D017353:Gene Deletion; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D051379:Mice; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D017354:Point Mutation; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D013438:Sulfhydryl Compounds / Q000378:metabolism; D013997:Time Factors; D019076:Transgenes; D020798:Two-Hybrid System Techniques; D025801:Ubiquitin / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism
D004952:Esters; D013438:Sulfhydryl Compounds; D025801:Ubiquitin; C471639:UBE3A protein, human; D044767:Ubiquitin-Protein Ligases; D005982:Glutathione Transferase