In vivo assembly of plasmid-expressed ribosomal protein S7 ofThermus thermophilusintoEscherichia coliribosomes and conditions of its overexpression
| Autor: | Andrey V. Karginov, Alexei M. Kopylov, V. A. Spiridonova, Olga A. Karginova |
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| Rok vydání: | 1995 |
| Předmět: |
Ribosomal Proteins
Overexpression Recombinant Fusion Proteins Genetic Vectors Molecular Sequence Data Biophysics Gene Expression Biology medicine.disease_cause Biochemistry Ribosome law.invention FLAG-tag Structural Biology Ribosomal protein law Escherichia coli Genetics medicine Assembly (in vivo) Amino Acid Sequence Molecular Biology Peptide sequence Base Sequence Thermus thermophilus Cell Biology Ribosomal protein S7 biology.organism_classification Fusion protein Molecular biology Genes Bacterial Recombinant DNA Ribosomes |
| Zdroj: | FEBS Letters. 369:158-160 |
| ISSN: | 0014-5793 |
| DOI: | 10.1016/0014-5793(95)00730-w |
| Popis: | Researchers still have great difficulty in isolating individual ribosomal proteins from the ribosome in quantities high enough for structural research. To this end, when studying protein S7, we created an E. coli overproducer of the recombinant protein S7 of Thermus thermophilus. The vector for expression was pQE-32 having a strong promoter of E. coli phage T5 and six triplets of His at the 5′-end. This N-terminal six His tag of the fusion protein is responsible for binding to Ni-NTA-resin and allows purifying the protein in one step. The yield of the recombinant protein was 20% and more of the total cellular proteins. In addition we have shown that the recombinant thermophilic protein is incorporated in vivo into the ribosome of E. coli despite the fact that these proteins (thermophilic and mesophilic) have a rather low homology, only 52%. This fact provides a base for the system to study functions of individual proteins. |
| Databáze: | OpenAIRE |
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