Source:http://linkedlifedata.com/resource/pubmed/id/15133025
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| Predicate | Object |
|---|---|
| rdf:type | |
| lifeskim:mentions | |
| pubmed:issue |
31
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| pubmed:dateCreated |
2004-7-26
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| pubmed:abstractText |
Residues in conserved motifs (625)TGD, (676)FARXXPXXK, and (701)TGDGVND in domain P of sarcoplasmic reticulum Ca(2+)-ATPase, as well as in motifs (601)DPPR and (359)NQR(/K)MSV in the hinge segments connecting domains N and P, were examined by mutagenesis to assess their roles in nucleotide and Mg(2+) binding and stabilization of the Ca(2+)-activated transition state for phosphoryl transfer. In the absence of Mg(2+), mutations removing the charges of domain P residues Asp(627), Lys(684), Asp(703), and Asp(707) increased the affinity for ATP and 2',3'-O-(2,4,6-trinitrophenyl)-8-azidoadenosine 5'-triphosphate. These mutations, as well as Gly(626)--> Ala, were inhibitory for ATP binding in the presence of Mg(2+) and for tight binding of the beta,gamma-bidentate chromium(III) complex of ATP. The hinge mutations had pronounced, but variable, effects on ATP binding only in the presence of Mg(2+). The data demonstrate an unfavorable electrostatic environment for binding of negatively charged nucleotide in domain P and show that Mg(2+) is required to anchor the phosphoryl group of ATP at the phosphorylation site. Mutants Gly(626) --> Ala, Lys(684) --> Met, Asp(703) --> Ala/Ser/Cys, and mutants with alteration to Asp(707) exhibited very slow or negligible phosphorylation, making it possible to measure ATP binding in the pseudo-transition state attained in the presence of both Mg(2+) and Ca(2+). Under these conditions, ATP binding was almost completely blocked in Gly(626) --> Ala and occurred with 12- and 7-fold reduced affinities in Asp(703) --> Ala and Asp(707) --> Cys, respectively, relative to the situation in the presence of Mg(2+) without Ca(2+), whereas in Lys(684) --> Met and Asp(707) --> Ser/Asn the affinity was enhanced 14- and 3-5-fold, respectively. Hence, Gly(626) and Asp(703) seem particularly critical for mediating entry into the transition state for phosphoryl transfer upon Ca(2+) binding at the transport sites.
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| pubmed:language |
eng
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| pubmed:journal | |
| pubmed:citationSubset |
IM
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| pubmed:chemical |
http://linkedlifedata.com/resource/pubmed/chemical/Adenosine Triphosphate,
http://linkedlifedata.com/resource/pubmed/chemical/Aspartic Acid,
http://linkedlifedata.com/resource/pubmed/chemical/Calcium,
http://linkedlifedata.com/resource/pubmed/chemical/Calcium-Transporting ATPases,
http://linkedlifedata.com/resource/pubmed/chemical/DNA, Complementary,
http://linkedlifedata.com/resource/pubmed/chemical/Glycine,
http://linkedlifedata.com/resource/pubmed/chemical/Magnesium,
http://linkedlifedata.com/resource/pubmed/chemical/Nucleotides,
http://linkedlifedata.com/resource/pubmed/chemical/Sarcoplasmic Reticulum...
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| pubmed:status |
MEDLINE
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| pubmed:month |
Jul
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| pubmed:issn |
0021-9258
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| pubmed:author | |
| pubmed:issnType |
Print
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| pubmed:day |
30
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| pubmed:volume |
279
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| pubmed:owner |
NLM
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| pubmed:authorsComplete |
Y
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| pubmed:pagination |
32515-23
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| pubmed:dateRevised |
2010-11-18
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| pubmed:meshHeading |
pubmed-meshheading:15133025-Adenosine Triphosphate,
pubmed-meshheading:15133025-Amino Acid Motifs,
pubmed-meshheading:15133025-Animals,
pubmed-meshheading:15133025-Aspartic Acid,
pubmed-meshheading:15133025-Biological Transport,
pubmed-meshheading:15133025-COS Cells,
pubmed-meshheading:15133025-Calcium,
pubmed-meshheading:15133025-Calcium-Transporting ATPases,
pubmed-meshheading:15133025-Catalysis,
pubmed-meshheading:15133025-DNA, Complementary,
pubmed-meshheading:15133025-Dose-Response Relationship, Drug,
pubmed-meshheading:15133025-Glycine,
pubmed-meshheading:15133025-Kinetics,
pubmed-meshheading:15133025-Magnesium,
pubmed-meshheading:15133025-Models, Molecular,
pubmed-meshheading:15133025-Mutagenesis,
pubmed-meshheading:15133025-Mutation,
pubmed-meshheading:15133025-Nucleotides,
pubmed-meshheading:15133025-Phosphorylation,
pubmed-meshheading:15133025-Protein Binding,
pubmed-meshheading:15133025-Protein Structure, Tertiary,
pubmed-meshheading:15133025-Rabbits,
pubmed-meshheading:15133025-Sarcoplasmic Reticulum,
pubmed-meshheading:15133025-Sarcoplasmic Reticulum Calcium-Transporting ATPases,
pubmed-meshheading:15133025-Time Factors
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| pubmed:year |
2004
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| pubmed:articleTitle |
Roles of conserved P domain residues and Mg2+ in ATP binding in the ground and Ca2+-activated states of sarcoplasmic reticulum Ca2+-ATPase.
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| pubmed:affiliation |
Chemical Pathology, Department of Clinical Laboratory Sciences, Faculty of Health Sciences, University of Cape Town, and National Health Laboratory Service, Groote Schuur Hospital, Cape Town 7925, South Africa. davidmci@chempath.uct.ac.za
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| pubmed:publicationType |
Journal Article,
Research Support, Non-U.S. Gov't
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