Residues interacting with serine-174 and alanine-295 in the β-subunit of Escherichia coli H+-ATP synthase: Possible ternary structure of the center region of the subunit
| Autor: | Yoshihisa Ishihara, Hiroshi Kanazawa, Junji Miki, Takashi Mano, Takato Noumi |
|---|---|
| Rok vydání: | 1994 |
| Předmět: |
ATPase
Protein subunit Molecular Sequence Data Biophysics Reversion medicine.disease_cause Biochemistry Serine Escherichia coli medicine Point Mutation Alanine chemistry.chemical_classification Mutation Binding Sites Base Sequence Molecular Structure biology Cell Biology Molecular biology Proton-Translocating ATPases Enzyme chemistry biology.protein |
| Zdroj: | Biochimica et Biophysica Acta (BBA) - Bioenergetics. 1187:67-72 |
| ISSN: | 0005-2728 |
| DOI: | 10.1016/0005-2728(94)90167-8 |
| Popis: | The mutation of serine-174 to phenylalanine that causes a defect in the Escherichia coli F 1 -ATPase β-subunit is suppressed by further mutations; Gly-149 to Ser, Ala-295 to Thr, Ala-295 to Pro, or Leu-400 to Gln (Miki, J., Fujiwara, K., Tsuda, M., Tsuchiya, T. and Kanazawa, H. (1990) J. Biol. Chem. 265, 21567–21572). We analyzed the effects of these second site mutations and of a newly identified Asn-158 to Tyr mutation on the activities of the ATPase without the original Ser-174 to Phe mutation. The β-subunit with each amino acid replacement was expressed in the mutant strain JP17, which does not have a β-subunit. Cells transformed with the plasmid carrying Ala-295 to Pro mutation alone did not grow on minimal medium agar supplemented with succinate as the sole carbon source, and showed 3% of the wild-type ATPase activity, suggesting that this mutation caused structural alterations affecting the catalytic function of the enzyme. Conversely transformants with other mutations grew well and had higher ATPase activities, suggesting that these mutations did not cause extensive structural alterations. From the transformants with the plasmid carrying the Ala-295 to Pro mutation, seven revertants capable of cell growth on succinate plates were isolated and reversion mutations were identified at residues 140, 159, 166, 171, 172 and 184 of the β-subunits. The results suggested that Ser-174 and Ala-295 do not necessarily interact directly, but that the regions including these suppression mutation sites close to Ser-174, and Ala-295 interact with each other for the proper functioning of the ATPase. The ternary structure of the region surrounded by the residues which were identified as the reversion mutation sites for Ser-174 to Phe and Ala-295 to Pro mutations is important for the catalytic function of this enzyme. |
| Databáze: | OpenAIRE |
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