Drosophila proteasome Dm25 subunit substitutes the mouse MC3 subunit in hybrid proteasomes. The N-terminal domain is essential for subunit incorporation.
The proteasome is a multisubunit 20 S proteinase complex involved in ubiquitin-dependent and -independent intracellular protein metabolism. Individual subunits of the alpha- and beta-type share extensive sequence homology and are encoded as members of two related and evolutionarily conserved gene families. Due to the lack of viable deletion mutants of essential alpha-type proteasome subunits in higher eukaryotes, an identification and analysis of potentially homologous subunits of different species was so far not possible. It is shown here that the novel Drosophila alpha-type Dm25 subunit can be incorporated into mouse proteasomes of stably transfected NIH 3T3 cells. The Dm25 subunit is able to substitute the mouse MC3 alpha-type subunit in proteasomes, indicating a high structural and possibly also functional homology of the two subunits. In contrast and pointing at the importance of the slightly hydrophobic N-terminal region stabile expression of a Dm25 subunit, which is truncated at its N terminus and lacks PROS box I, results in a subunit which cannot be incorporated into mouse proteasomes. The ability to form hybrid proteasomes involving essential nondeletable subunits now opens the possibility for structural and also functional analysis of such subunits by mutagenesis in higher eukaryotes.
Seelig
A
A
Center of Molecular Biology, University of Heidelberg, Federal Republic of Germany.
Multhaup
G
G
Pesold-Hurt
B
B
Beyreuther
K
K
Kloetzel
P M
PM
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003546:Cysteine Endopeptidases / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D004330:Drosophila / Q000201:enzymology*; D046911:Macromolecular Substances; D051379:Mice; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000096:biosynthesis / Q000302:isolation & purification / Q000378:metabolism*; D009838:Oligodeoxyribonucleotides; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D046988:Proteasome Endopeptidase Complex; D055503:Protein Multimerization; D015183:Restriction Mapping; D017386:Sequence Homology, Amino Acid; D014162:Transfection
D046911:Macromolecular Substances; D009097:Multienzyme Complexes; D009838:Oligodeoxyribonucleotides; D010446:Peptide Fragments; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex