| Autor: |
Pirch, Torsten, Quick, Matthias, Nietschke, Monika, Langkamp, Markus, Jung, Heinrich |
| Zdroj: |
Journal of Biological Chemistry; March 2002, Vol. 277 Issue: 11 p8790-8796, 7p |
| Abstrakt: |
To elucidate the functional importance of transmembrane domain II in the Na+/proline transporter (PutP) of Escherichia coliwe analyzed the effect of replacing Ser-54 through Gly-58. Substitution of Asp-55 or Met-56 dramatically reduces the apparent affinity for Na+and Li+in a cation-dependent manner. Conversely, Cys in place of Gly-58 significantly reduces only the apparent proline affinity while substitution of Ser-57 results in a dramatic reduction of the apparent proline andcation affinities. Interestingly, upon increasing the proline concentration the apparent Na+affinity of Ser-57 replacement mutants converges toward the wild-type value, indicating a close cooperativity between cation and substrate site(s). This notion is supported by the fact that Na+-stimulated site-specific fluorescence labeling of a single Cys at position 57 is completely reversed by the addition of proline. Similar results are obtained upon labeling of a Cys at position 54 or 58. Taken together, these results indicate that Asp-55 and Met-56 are located at or close to the ion-binding site while Ser-54, Ser-57, and Gly-58 may be close to the proline translocation pathway. In addition, the data prod at an involvement of the latter residues in ligand-induced conformational dynamics that are crucial for cation-coupled transport. |
| Databáze: |
Supplemental Index |
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