Purification, Cloning, and Characterization of the 16S RNA m5C967 Methyltransferase fromEscherichia coli
| Autor: | Marek Sochacki, James Ofengand, Joan S. Tscherne, Paul Popienick, Hanspeter Michel, Kelvin Nurse |
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| Rok vydání: | 1999 |
| Předmět: |
Molecular Sequence Data
Gene Expression Biology medicine.disease_cause Biochemistry Substrate Specificity law.invention Gene product Cytosine Open Reading Frames law RNA Ribosomal 16S Escherichia coli medicine Magnesium 30S Amino Acid Sequence Cloning Molecular Gene Methyltransferases DNA Methylation Ribosomal RNA Molecular biology Recombinant Proteins Open reading frame Genes Bacterial 5-Methylcytosine Recombinant DNA DNMT1 |
| Zdroj: | Biochemistry. 38:1884-1892 |
| ISSN: | 1520-4995 0006-2960 |
| Popis: | The methyltransferase that forms m5C967 in Escherichia coli small subunit ribosomal RNA has been purified, cloned, and characterized. The gene was identified from the N-terminal sequence of the purified enzyme. The gene is a fusion of two open reading frames, fmu and fmv, previously believed to be distinct due to a DNA sequencing error. The gene, here named rsmB, encodes a 429-amino acid protein that has a number of homologues in prokaryotes, Archaea, and eukaryotes. C-Terminal sequencing of the overexpressed and affinity-purified protein by mass spectrometry methods verified the sequence expected for the gene product. The recombinant protein exhibited the same specificity as the previously described native enzyme; that is, it formed only m5C and only at position 967. C1407, which is also m5C in natural 16S RNA, was not methylated. In vitro, the enzyme only recognized free 16S RNA. 30S ribosomal subunits were not a substrate. There was no requirement for added magnesium, suggesting that extensive secondary or tertiary structure in the RNA substrate may not be a requirement for recognition. |
| Databáze: | OpenAIRE |
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