Genetic, enzymatic, and structural analyses of phenylalanyl-tRNA synthetase from Thermococcus kodakaraensis KOD1
| Autor: | Kenzo Fujimoto, Kentaro Shiraki, Masao Tsuji, Yoshiteru Hashimoto, Masahiro Takagi, Tadayuki Imanaka, Shinsuke Fujiwara |
|---|---|
| Rok vydání: | 2003 |
| Předmět: |
Protein Folding
phenylalanyl-tRNA synthetase Transcription Genetic hyperthermophilic protein TATA box Molecular Sequence Data Static Electricity Oligonucleotides Biology medicine.disease_cause Biochemistry Open Reading Frames medicine Amino Acid Sequence Cloning Molecular Molecular Biology Escherichia coli Phylogeny Thermostability chemistry.chemical_classification heterooligomeric protein Base Sequence Sequence Homology Amino Acid Circular Dichroism Temperature General Medicine biology.organism_classification Molecular biology Protein tertiary structure Recombinant Proteins Amino acid Protein Structure Tertiary Thermococcus Open reading frame Isoelectric point chemistry electrostatic interaction Electrophoresis Polyacrylamide Gel Phenylalanine-tRNA Ligase Isoelectric Focusing |
| Zdroj: | Journal of biochemistry. 134(4) |
| ISSN: | 0021-924X |
| Popis: | Phenylalanyl-tRNA synthetase from the hyperthermophilic archaeon Thermococcus kodakaraensis KOD1 (Tk-PheRS) was cloned. The open reading frames for both the alpha-subunit (Tk-pheRSA) and beta-subunit (Tk-pheRSB) genes were 1,503 bp (501 amino acids) and 1,722 bp (574 amino acids), respectively. Tk-pheRSB located 879 bp downstream from Tk-pheRSA with a putative TATA box, suggesting that these two subunits are transcribed and regulated independently in KOD1 cells. Tk-PheRS and its respective subunits were expressed in Escherichia coli cells and the proteins were purified. Tk-PheRS showed an optimum enzymatic activity at around 95 degrees C and retained its tertiary structure at 98 degrees C. The estimated isoelectric point (pI) for the alpha-subunit is 9.4 and that for the beta-subunit is 4.6, the largest difference among the 12 kinds of PheRSs reported. The considerable thermostability of Tk-PheRS may be responsible for the electrostatic interaction between the alpha- and beta-subunits. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|