The phosphorylated ribosomal protein S7 in Tetrahymena is homologous with mammalian S4 and the phosphorylated residues are located in the C-terminal region. Structural characterization of proteins separated by two-dimensional polyacrylamide gel electrophoresis.
A single basic ribosomal protein, protein S7, can be multiply phosphorylated in the ciliated protozoan Tetrahymena. Induction of phosphorylation is highly regulated, and the phosphorylation proceeds in a strictly sequential manner. The first site to be phosphorylated is a serine residue and the second a threonine. In this paper we report the complete primary structure of Tetrahymena thermophila ribosomal protein S7 including identification of the phosphorylated serine and threonine residues. Most of the sequence information was obtained from peptides generated by in situ digestion of S7 in two-dimensional gels using an approach that combined traditional protein chemistry with mass spectrometry. T. thermophila ribosomal protein S7 has a molecular mass of 29,459 Da and contains 259 amino acid residues. Phosphorylation takes place on Ser258 and Thr248 in the C-terminal region of the protein. Alignment of T. thermophila ribosomal protein S7 with known ribosomal proteins yielded the surprising result that T. thermophila S7 is homologous, not with mammalian ribosomal protein S6, but with mammalian ribosomal protein S4. These findings clearly distinguish the pattern of phosphorylation of ribosomal proteins in Tetrahymena from all other eukaryotes analyzed to date.
Palm
L
L
Department of Molecular Biology, Odense University, Denmark.
Andersen
J
J
Rahbek-Nielsen
H
H
Hansen
T S
TS
Kristiansen
K
K
Højrup
P
P
D000595:Amino Acid Sequence; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D003488:Cyanogen Bromide; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D013058:Mass Spectrometry; D017017:Methanococcus; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D010766:Phosphorylation; D010768:Phosphoserine / Q000032:analysis; D010769:Phosphothreonine / Q000032:analysis; D051381:Rats; D012269:Ribosomal Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D012441:Saccharomyces cerevisiae; D017386:Sequence Homology, Amino Acid; D016808:Tetrahymena thermophila / Q000254:growth & development / Q000378:metabolism*; D014357:Trypsin
D010446:Peptide Fragments; D012269:Ribosomal Proteins; C037442:ribosomal protein S7; D010769:Phosphothreonine; D010768:Phosphoserine; D014357:Trypsin; D003488:Cyanogen Bromide