A Multiubiquitin Chain Is Confined to Specific Lysine in a Targeted Short-Lived Protein

TLDR

Ubiquitin-dependent degradation of β‑galactosidase is preceded by the attachment of an ordered, branched multi‑ubiquitin chain linked via Gly76–Lys48, and specific lysine residues in β‑galactosidase serve as the amino‑terminal degradation signal. The multi‑ubiquitin chain attaches to one of two specific lysine residues in β‑galactosidase, and mutating ubiquitin Lys48 demonstrates that this chain is essential for protein degradation.

Abstract

The ubiquitin-dependent degradation of a test protein β-galactosidase (βgal) is preceded by ubiquitination of βgal. The many (from 1 to more than 20) ubiquitin moieties attached to a molecule of βgal occur as an ordered chain of branched ubiquitin-ubiquitin conjugates in which the carboxyl-terminal Gly 76 of one ubiquitin is joined to the internal Lys 48 of an adjacent ubiquitin. This multiubiquitin chain is linked to one of two specific Lys residues in βgal. These same Lys residues have been identified by molecular genetic analysis as components of the amino-terminal degradation signal in βgal. The experiments with ubiquitin mutated at its Lys 48 residue indicate that the multiubiquitin chain in a targeted protein is essential for the degradation of the protein.