Localization of Saccharomyces cerevisiae ribosomal protein L16 on the surface of 60 S ribosomal subunits by immunoelectron microscopy.
Antibodies raised against a trpE-L16 fusion protein expressed in Escherichia coli were used to examine immunological relatedness between Saccharomyces cerevisiae ribosomal protein L16 and ribosomal proteins from eubacteria, halobacteria, methanogens, eocytes, and other eukaryotes. Homologues of L16 also were identified by searches of sequence data bases. Among the bacterial proteins that are immunologically related and similar in sequence to L16 are ribosomal proteins that bind 5 S rRNA. L16 protein fused near its carboxyl terminus to E. coli beta-galactosidase could assemble into functional yeast 60 S ribosomal subunits. The RPL16A-lacZ gene fusion partially complemented the slow growth or lethality of mutants containing null alleles of one or both RPL16 genes, respectively. L16-beta-galactosidase fusion protein cosedimented with ribosomes and polyribosomes, and remained associated with high salt-washed ribosomes. Monoclonal antibodies against beta-galactosidase were used to map the location of L16-beta-galactosidase on the surface of the 60 S subunit by immunoelectron microscopy. L16 was localized near the top surface of the central protuberance, where the 60 S subunit potentially contacts the 40 S subunit. This is similar to the location of the bacterial homologues of L16 in 50 S ribosomal subunits.
Tsay
Y F
YF
Department of Biological Sciences, Carnegie Mellon University, Pittsburgh, Pennsylvania 15213.
Shankweiler
G
G
Lake
J
J
Woolford
J L
JL
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D001419:Bacteria / Q000378:metabolism; D007425:Intracellular Membranes / Q000737:chemistry / Q000648:ultrastructure; D016253:Microscopy, Immunoelectron; D008969:Molecular Sequence Data; D012329:RNA, Bacterial / Q000378:metabolism; D012331:RNA, Fungal / Q000378:metabolism; D012341:RNA, Ribosomal, 5S / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012269:Ribosomal Proteins / Q000032:analysis* / Q000276:immunology; D012270:Ribosomes / Q000737:chemistry* / Q000648:ultrastructure; D012441:Saccharomyces cerevisiae / Q000737:chemistry* / Q000276:immunology; D001616:beta-Galactosidase / Q000276:immunology / Q000378:metabolism
D000911:Antibodies, Monoclonal; D012329:RNA, Bacterial; D012331:RNA, Fungal; D012341:RNA, Ribosomal, 5S; D016601:RNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D012269:Ribosomal Proteins; C027937:ribosomal protein L16; D001616:beta-Galactosidase