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PMID:19922410
| Citation |
Karasawa, A, Mitsui, K, Matsushita, M and Kanazawa, H (2010) Intermolecular cross-linking of monomers in Helicobacter pylori Na+/H+ antiporter NhaA at the dimer interface inhibits antiporter activity. Biochem. J. 426:99-108 |
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| Abstract |
We have previously shown that HPNhaA (Helicobacter pylori Na+/H+ antiporter) forms an oligomer in a native membrane of Escherichia coli, and conformational changes of oligomer occur between monomers of the oligomer during ion transport. In the present study, we use Blue-native PAGE to show that HPNhaA forms a dimer. Cysteine-scanning mutagenesis of residues 55-61 in a putative beta-sheet region of loop1 and subsequent functional analyses revealed that the Q58C mutation resulted in an intermolecular disulfide bond. G56C, I59C and G60C were found to be cross-linked by bifunctional cross-linkers. Furthermore, the Q58E mutant did not form a dimer, possibly due to electrostatic repulsion between monomers. These results imply that Gln-58 and the flanking sequence in the putative beta-sheet of the monomer are located close to the identical residues in the dimer. The Q58C mutant of NhaA was almost inactive under non-reducing conditions, and activity was restored under reducing conditions. This result showed that cross-linking at the dimer interface reduces transporter activity by interfering with the flexible association between the monomers. A mutant HPNhaA protein with three amino acid substitutions at residues 57-59 did not form a dimer, and yet was active, indicating that the monomer is functional. |
| Links |
PubMed Online version:10.1042/BJ20091339 |
| Keywords |
Bacterial Proteins/chemistry; Bacterial Proteins/metabolism; Helicobacter pylori/metabolism; Mutagenesis, Site-Directed; Protein Multimerization; Protein Structure, Tertiary; Sodium-Hydrogen Antiporter/chemistry; Sodium-Hydrogen Antiporter/metabolism |
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Significance
Annotations
| Gene product | Qualifier | GO Term | Evidence Code | with/from | Aspect | Extension | Notes | Status |
|---|---|---|---|---|---|---|---|---|
| GO:0006811 : ion transport |
ECO:0000315: |
P |
Figure 1, 2, and 3A all support the idea that Q58C is a mutant. This is because it creates dimers as the paper suggested it would. |
complete | ||||
See also
References
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