cDNA cloning and functional analysis of p28 (Nas6p) and p40.5 (Nas7p), two novel regulatory subunits of the 26S proteasome.
We employed cDNA cloning to deduce the complete primary structures of p28 and p40.5, two novel subunits of PA700 (also called 19S complex), a 700 kDa multisubunit regulatory complex of the human 26S proteasome. These polypeptides consisted of 226 and 376 amino acids with calculated molecular masses of 24428 Da and 42945 Da, and isoelectric points of 5. 68 and 5.46, respectively. Intriguingly, p28 contained five conserved motifs known as 'ankyrin repeats', implying that this subunit may contribute to interaction of the 26S proteasome with other protein(s). Computer-assisted homology analysis revealed high sequence similarities of p28 and p40.5 with yeast proteins, termed Nas6p and Nas7p (non-ATPase subunits 6 and 7), respectively, whose functions are as yet unknown. Disruption of these yeast genes, NAS6 and NAS7, had no effect on cell viability, indicating that neither of the two subunits is essential for proliferation of yeast cells. However, the NAS7, but not NAS6, disruptant cells caused high sensitivity to heat stress, being unable to proliferate at 37 degreesC.
Hori
T
T
The Tokyo Metropolitan Institute of Medical Science, and CREST, Japan Science and Technology Corporation (JST), 3-18-22 Honkomagome, Bunkyo-ku, Tokyo 113-8613, Japan.
Kato
S
S
Saeki
M
M
DeMartino
G N
GN
Slaughter
C A
CA
Takeuchi
J
J
Toh-e
A
A
Tanaka
K
K
D000251:Adenosine Triphosphatases / Q000235:genetics; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001483:Base Sequence; D002417:Cattle; D002455:Cell Division / Q000235:genetics / Q000502:physiology; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics*; D010450:Endopeptidases; D005656:Fungal Proteins / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic; D005800:Genes, Fungal / Q000235:genetics; D006358:Hot Temperature; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010447:Peptide Hydrolases / Q000737:chemistry / Q000235:genetics*; D046988:Proteasome Endopeptidase Complex; D011506:Proteins / Q000737:chemistry / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000201:enzymology / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D014018:Tissue Distribution; D014407:Tumor Cells, Cultured
D000596:Amino Acids; D018076:DNA, Complementary; D005656:Fungal Proteins; C114551:NAS6 protein, S cerevisiae; D011506:Proteins; D012333:RNA, Messenger; D029701:Saccharomyces cerevisiae Proteins; D010450:Endopeptidases; D010447:Peptide Hydrolases; D046988:Proteasome Endopeptidase Complex; C061553:ATP dependent 26S protease; C119011:26S proteasome non-ATPase regulatory subunit 13; D000251:Adenosine Triphosphatases