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Calcium-induced conformational changes in the regulatory domain of the human mitochondrial ATP-Mg/Pi carrier.

Harborne SP, Ruprecht JJ, Kunji ER - Biochim. Biophys. Acta (2015)

Bottom Line: Careful analysis by SEC confirmed that although the regulatory domain crystallised as dimers, full-length ATP-Mg/Pi carrier is monomeric.Detailed bioinformatics analyses of different EF-hand states indicate that upon release of calcium, EF-hands close, meaning that the regulatory domain would release the amphipathic α-helix.We propose a mechanism for ATP-Mg/Pi carriers in which the amphipathic α-helix becomes mobile upon release of calcium and could block the transport of substrates across the mitochondrial inner membrane.

View Article: PubMed Central - PubMed

Affiliation: The Medical Research Council, Mitochondrial Biology Unit, Cambridge Biomedical Campus, Wellcome Trust/MRC Building, Hills Road, Cambridge CB2 0XY, UK.

No MeSH data available.


A model for the calcium regulation of HsAPC-1 RD.A) Three models for calcium-free HsAPC-1 RD based on centrin (model-1), calmodulin (model-2) and calcium-dependent protein kinase (model-3) were superposed onto the calcium-bound HsAPC-1 RD structure and coloured cyan, magenta and green respectively. The calcium-bound HsAPC-1 structure is coloured by RMSD difference from the calcium-free models as in the key. B) A close up view of hydrophobic pocket 2 into which the amphipathic α-helix (red) is bound. The calcium-bound HsAPC-1 RD structure is shown as a surface representation and coloured blue, whereas the superposed calcium-free models are represented as cartoon (coloured as in panel A). C) The proposed regulatory mechanism between calcium-bound state of HsAPC-1 where the carrier is active and the calcium-free state of HsAPC-1 where the carrier is inactive. The calcium-free model presented in panel C is based on model 2. α-helices are represented as cylinders in panel C. Calcium ions are presented as lime green spheres.
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f0025: A model for the calcium regulation of HsAPC-1 RD.A) Three models for calcium-free HsAPC-1 RD based on centrin (model-1), calmodulin (model-2) and calcium-dependent protein kinase (model-3) were superposed onto the calcium-bound HsAPC-1 RD structure and coloured cyan, magenta and green respectively. The calcium-bound HsAPC-1 structure is coloured by RMSD difference from the calcium-free models as in the key. B) A close up view of hydrophobic pocket 2 into which the amphipathic α-helix (red) is bound. The calcium-bound HsAPC-1 RD structure is shown as a surface representation and coloured blue, whereas the superposed calcium-free models are represented as cartoon (coloured as in panel A). C) The proposed regulatory mechanism between calcium-bound state of HsAPC-1 where the carrier is active and the calcium-free state of HsAPC-1 where the carrier is inactive. The calcium-free model presented in panel C is based on model 2. α-helices are represented as cylinders in panel C. Calcium ions are presented as lime green spheres.

Mentions: Although the three resulting models had some structural differences from one another, the general motion that they described was consistent (Fig. 5A). All calcium-free models show that α-helices 6 and 7 adopt a position that would prevent the binding of the amphipathic α-helix to hydrophobic pocket 2 (Fig. 5B). Therefore, in the absence of calcium, the hydrophobic pockets of HsAPC-1 RD close, and as a consequence the amphipathic α-helix is excluded (Supplementary video 1). All three models agree with this mechanism, but model 2 is preferred, as it represents the median of calcium-free EF-hand structures (Fig. 4G). In support of this mechanism, the isolated APC regulatory domain becomes more dynamic in the absence of calcium, particularly in the region of the amphipathic α-helix as demonstrated by nuclear Overhauser effect experiments [16]. Furthermore, we demonstrate here using SEC that under calcium-free condition the hydrodynamic radius of the isolated APC regulatory domain is increased in comparison with the calcium-bound state (Table 4), consistent with the exclusion of the amphipathic α-helix from the pocket.


Calcium-induced conformational changes in the regulatory domain of the human mitochondrial ATP-Mg/Pi carrier.

Harborne SP, Ruprecht JJ, Kunji ER - Biochim. Biophys. Acta (2015)

A model for the calcium regulation of HsAPC-1 RD.A) Three models for calcium-free HsAPC-1 RD based on centrin (model-1), calmodulin (model-2) and calcium-dependent protein kinase (model-3) were superposed onto the calcium-bound HsAPC-1 RD structure and coloured cyan, magenta and green respectively. The calcium-bound HsAPC-1 structure is coloured by RMSD difference from the calcium-free models as in the key. B) A close up view of hydrophobic pocket 2 into which the amphipathic α-helix (red) is bound. The calcium-bound HsAPC-1 RD structure is shown as a surface representation and coloured blue, whereas the superposed calcium-free models are represented as cartoon (coloured as in panel A). C) The proposed regulatory mechanism between calcium-bound state of HsAPC-1 where the carrier is active and the calcium-free state of HsAPC-1 where the carrier is inactive. The calcium-free model presented in panel C is based on model 2. α-helices are represented as cylinders in panel C. Calcium ions are presented as lime green spheres.
© Copyright Policy - CC BY
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4562336&req=5

f0025: A model for the calcium regulation of HsAPC-1 RD.A) Three models for calcium-free HsAPC-1 RD based on centrin (model-1), calmodulin (model-2) and calcium-dependent protein kinase (model-3) were superposed onto the calcium-bound HsAPC-1 RD structure and coloured cyan, magenta and green respectively. The calcium-bound HsAPC-1 structure is coloured by RMSD difference from the calcium-free models as in the key. B) A close up view of hydrophobic pocket 2 into which the amphipathic α-helix (red) is bound. The calcium-bound HsAPC-1 RD structure is shown as a surface representation and coloured blue, whereas the superposed calcium-free models are represented as cartoon (coloured as in panel A). C) The proposed regulatory mechanism between calcium-bound state of HsAPC-1 where the carrier is active and the calcium-free state of HsAPC-1 where the carrier is inactive. The calcium-free model presented in panel C is based on model 2. α-helices are represented as cylinders in panel C. Calcium ions are presented as lime green spheres.
Mentions: Although the three resulting models had some structural differences from one another, the general motion that they described was consistent (Fig. 5A). All calcium-free models show that α-helices 6 and 7 adopt a position that would prevent the binding of the amphipathic α-helix to hydrophobic pocket 2 (Fig. 5B). Therefore, in the absence of calcium, the hydrophobic pockets of HsAPC-1 RD close, and as a consequence the amphipathic α-helix is excluded (Supplementary video 1). All three models agree with this mechanism, but model 2 is preferred, as it represents the median of calcium-free EF-hand structures (Fig. 4G). In support of this mechanism, the isolated APC regulatory domain becomes more dynamic in the absence of calcium, particularly in the region of the amphipathic α-helix as demonstrated by nuclear Overhauser effect experiments [16]. Furthermore, we demonstrate here using SEC that under calcium-free condition the hydrodynamic radius of the isolated APC regulatory domain is increased in comparison with the calcium-bound state (Table 4), consistent with the exclusion of the amphipathic α-helix from the pocket.

Bottom Line: Careful analysis by SEC confirmed that although the regulatory domain crystallised as dimers, full-length ATP-Mg/Pi carrier is monomeric.Detailed bioinformatics analyses of different EF-hand states indicate that upon release of calcium, EF-hands close, meaning that the regulatory domain would release the amphipathic α-helix.We propose a mechanism for ATP-Mg/Pi carriers in which the amphipathic α-helix becomes mobile upon release of calcium and could block the transport of substrates across the mitochondrial inner membrane.

View Article: PubMed Central - PubMed

Affiliation: The Medical Research Council, Mitochondrial Biology Unit, Cambridge Biomedical Campus, Wellcome Trust/MRC Building, Hills Road, Cambridge CB2 0XY, UK.

No MeSH data available.