Inhibition of ubiquitin-dependent proteolysis by a synthetic glycine-alanine repeat peptide that mimics an inhibitory viral sequence.
The glycine-alanine repeat (GAr) of the Epstein-Barr virus nuclear antigen-1 is a cis-acting transferable element that inhibits ubiquitin/proteasome-dependent proteolysis in vitro and in vivo. We have here examined the effect of a synthetic 20-mer GAr oligopeptide on the degradation of iodinated or biotin labeled lysozyme in a rabbit reticulocyte lysates in vitro assay. Micromolar concentrations of the GA-20 peptide inhibited the hydrolysis of lysozyme without significant effect on ubiquitination. Addition of the peptide did not inhibit the hydrolysis of fluorogenic substrate by purified proteasomes and did not affect the ubiquitination of lysozyme. An excess of the peptide failed to compete for binding of a synthetic tetra-ubiquitin complex to the S5a ubiquitin-binding subunit of the 19S regulator, confirming that the GAr does not block the access of ubiquitinated substrates to the proteasome. Our data suggest that the GAr may act by destabilizing the interaction of ubiquitinated substrates with the proteasome and promote the premature release of the substrate.
Leonchiks
Ainars
A
Microbiology and Tumor Biology Center, Karolinska Institutet, S-171 77 Stockholm, Sweden.
Stavropoulou
Vaya
V
Sharipo
Anatoly
A
Masucci
Maria G
MG
D000409:Alanine; D000595:Amino Acid Sequence; D000818:Animals; D001710:Biotin; D002352:Carrier Proteins / Q000037:antagonists & inhibitors; D003546:Cysteine Endopeptidases; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D019309:Epstein-Barr Virus Nuclear Antigens / Q000494:pharmacology*; D005998:Glycine; D004854:Herpesvirus 4, Human; D007457:Iodine Radioisotopes; D007553:Isotope Labeling; D018716:Molecular Mimicry; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors*; D009113:Muramidase / Q000037:antagonists & inhibitors*; D009842:Oligopeptides / Q000494:pharmacology*; D010455:Peptides / Q000494:pharmacology*; D011108:Polymers; D046988:Proteasome Endopeptidase Complex; D011817:Rabbits; D014452:Ubiquitins / Q000378:metabolism*
D002352:Carrier Proteins; D004791:Enzyme Inhibitors; D019309:Epstein-Barr Virus Nuclear Antigens; D007457:Iodine Radioisotopes; D009097:Multienzyme Complexes; D009842:Oligopeptides; D010455:Peptides; D011108:Polymers; D014452:Ubiquitins; C034500:poly(alanylglycine); D001710:Biotin; C421864:hen egg lysozyme; D009113:Muramidase; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; C417029:EBV-encoded nuclear antigen 1; D000409:Alanine; D005998:Glycine