Mass spectrometric analysis of 40 S ribosomal proteins from Rat-1 fibroblasts.
Although sequences of most mammalian ribosomal proteins are available, little is known about the post-translational processing of ribosomal proteins. To examine their post-translational modifications, 40 S subunit proteins purified from Rat-1 fibroblasts and their peptides were analyzed by liquid chromatography coupled with electrospray mass spectrometry. Of 41 proteins observed, 36 corresponded to the 32 rat 40 S ribosomal proteins with known sequences (S3, S5, S7, and S24 presented in two forms). The observed masses of S4, S6-S8, S13, S15a, S16, S17, S19, S27a, S29, and S30 matched those predicted. Sa, S3a, S5, S11, S15, S18, S20, S21, S24, S26-S28, and an S7 variant showed changes in mass that were consistent with N-terminal demethionylation and/or acetylation (S5 and S27 also appeared to be internally formylated and acetylated, respectively). S23 appeared to be internally hydroxylated or methylated. S2, S3, S9, S10, S12, S14, and S25 showed changes in mass inconsistent with known covalent modifications (+220, -75, +86, +56, -100, -117, and -103 Da, respectively), possibly representing novel post-translational modifications or allelic sequence variation. Five unidentified proteins (12,084, 13,706, 13,741, 13,884, and 34, 987 Da) were observed; for one, a sequence tag (PPGPPP), absent in any known ribosomal proteins, was determined, suggesting that it is a previously undescribed ribosome-associated protein. This study establishes a powerful method to rapidly analyze protein components of large biological complexes and their covalent modifications.
Howard Hughes Medical Institute, University of Colorado, Boulder, Colorado 80309, USA. louied@stripe.Colorado.edu
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Louie
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Resing
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Lewis
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D000595:Amino Acid Sequence; D000818:Animals; D002478:Cells, Cultured; D005347:Fibroblasts / Q000737:chemistry; D013058:Mass Spectrometry; D008666:Metalloendopeptidases / Q000378:metabolism; D008969:Molecular Sequence Data; D051381:Rats; D012269:Ribosomal Proteins / Q000737:chemistry*
D012269:Ribosomal Proteins; C041308:ribosomal protein S40; D008666:Metalloendopeptidases; C039830:peptidyl-Lys metalloendopeptidase