Identification and characterisation of the α and β subunits of succinyl CoA ligase of tomato
Despite the central importance of the TCA cycle in plant metabolism not all of the genes encoding its constituent enzymes have been functionally identified. In yeast, the heterodimeric protein succinyl CoA ligase is encoded for by two single-copy genes. Here we report the isolation of two tomato cDN...
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todo:paper_01674412_v59_n5_p781_StudartGuimaraes2023-10-03T15:04:42Z Identification and characterisation of the α and β subunits of succinyl CoA ligase of tomato Studart-Guimarães, C. Gibon, Y. Frankel, N. Wood, C.C. Zanor, M.I. Fernie, A.R. Carrari, F. Mitochondrial metabolism Saccharomyces cerevisiae Solanum lycopersicum Succinyl CoA ligase Tricarboxylic acid cycle Crops Cytology Enzymes Genes Genetic engineering Metabolism Yeast Mitochondrial metabolism Solanum licopersicum Succinyl CoA ligase Tricarboxylic acid cycle Enzyme kinetics protein subunit succinic acid succinyl coenzyme A synthetase amino acid sequence article chemistry cytology DNA sequence enzymology gene expression regulation genetic complementation genetics metabolism molecular cloning phylogeny protein tertiary structure protein transport sequence homology tomato Amino Acid Sequence Cloning, Molecular Gene Expression Regulation, Enzymologic Gene Expression Regulation, Plant Genetic Complementation Test Lycopersicon esculentum Phylogeny Protein Structure, Tertiary Protein Subunits Protein Transport Sequence Analysis, DNA Sequence Homology, Amino Acid Succinate-CoA Ligases Succinic Acid Cytology Enzymatic Activity Enzymes Genes Genetic Engineering Metabolism Saccharomyces Cerevisiae Tomatoes Lycopersicon esculentum Saccharomyces cerevisiae Solanum Despite the central importance of the TCA cycle in plant metabolism not all of the genes encoding its constituent enzymes have been functionally identified. In yeast, the heterodimeric protein succinyl CoA ligase is encoded for by two single-copy genes. Here we report the isolation of two tomato cDNAs coding for α- and one coding for the β-subunit of succinyl CoA ligase. These three cDNAs were used to complement the respective Saccharomyces cerevisiae mutants deficient in the α- and β-subunit, demonstrating that they encode functionally active polypeptides. The genes encoding for the subunits were expressed in all tissues, but most strongly in floral and leaf tissues, with equivalent expression of the two α-subunit genes being expressed to equivalent levels in all tissues. In all instances GFP fusion expression studies confirmed an expected mitochondrial location of the proteins encoded. Following the development of a novel assay to measure succinyl CoA ligase activity, in the direction of succinate formation, the evaluation of the maximal catalytic activities of the enzyme in a range of tissues revealed that these paralleled those of mRNA levels. We also utilized this assay to perform a preliminary characterisation of the regulatory properties of the enzyme suggesting allosteric control of this enzyme which may regulate flux through the TCA cycle in a manner consistent with its position therein. © Springer 2005. Fil:Frankel, N. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Carrari, F. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. JOUR info:eu-repo/semantics/openAccess http://creativecommons.org/licenses/by/2.5/ar http://hdl.handle.net/20.500.12110/paper_01674412_v59_n5_p781_StudartGuimaraes |
| institution |
Universidad de Buenos Aires |
| institution_str |
I-28 |
| repository_str |
R-134 |
| collection |
Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA) |
| topic |
Mitochondrial metabolism Saccharomyces cerevisiae Solanum lycopersicum Succinyl CoA ligase Tricarboxylic acid cycle Crops Cytology Enzymes Genes Genetic engineering Metabolism Yeast Mitochondrial metabolism Solanum licopersicum Succinyl CoA ligase Tricarboxylic acid cycle Enzyme kinetics protein subunit succinic acid succinyl coenzyme A synthetase amino acid sequence article chemistry cytology DNA sequence enzymology gene expression regulation genetic complementation genetics metabolism molecular cloning phylogeny protein tertiary structure protein transport sequence homology tomato Amino Acid Sequence Cloning, Molecular Gene Expression Regulation, Enzymologic Gene Expression Regulation, Plant Genetic Complementation Test Lycopersicon esculentum Phylogeny Protein Structure, Tertiary Protein Subunits Protein Transport Sequence Analysis, DNA Sequence Homology, Amino Acid Succinate-CoA Ligases Succinic Acid Cytology Enzymatic Activity Enzymes Genes Genetic Engineering Metabolism Saccharomyces Cerevisiae Tomatoes Lycopersicon esculentum Saccharomyces cerevisiae Solanum |
| spellingShingle |
Mitochondrial metabolism Saccharomyces cerevisiae Solanum lycopersicum Succinyl CoA ligase Tricarboxylic acid cycle Crops Cytology Enzymes Genes Genetic engineering Metabolism Yeast Mitochondrial metabolism Solanum licopersicum Succinyl CoA ligase Tricarboxylic acid cycle Enzyme kinetics protein subunit succinic acid succinyl coenzyme A synthetase amino acid sequence article chemistry cytology DNA sequence enzymology gene expression regulation genetic complementation genetics metabolism molecular cloning phylogeny protein tertiary structure protein transport sequence homology tomato Amino Acid Sequence Cloning, Molecular Gene Expression Regulation, Enzymologic Gene Expression Regulation, Plant Genetic Complementation Test Lycopersicon esculentum Phylogeny Protein Structure, Tertiary Protein Subunits Protein Transport Sequence Analysis, DNA Sequence Homology, Amino Acid Succinate-CoA Ligases Succinic Acid Cytology Enzymatic Activity Enzymes Genes Genetic Engineering Metabolism Saccharomyces Cerevisiae Tomatoes Lycopersicon esculentum Saccharomyces cerevisiae Solanum Studart-Guimarães, C. Gibon, Y. Frankel, N. Wood, C.C. Zanor, M.I. Fernie, A.R. Carrari, F. Identification and characterisation of the α and β subunits of succinyl CoA ligase of tomato |
| topic_facet |
Mitochondrial metabolism Saccharomyces cerevisiae Solanum lycopersicum Succinyl CoA ligase Tricarboxylic acid cycle Crops Cytology Enzymes Genes Genetic engineering Metabolism Yeast Mitochondrial metabolism Solanum licopersicum Succinyl CoA ligase Tricarboxylic acid cycle Enzyme kinetics protein subunit succinic acid succinyl coenzyme A synthetase amino acid sequence article chemistry cytology DNA sequence enzymology gene expression regulation genetic complementation genetics metabolism molecular cloning phylogeny protein tertiary structure protein transport sequence homology tomato Amino Acid Sequence Cloning, Molecular Gene Expression Regulation, Enzymologic Gene Expression Regulation, Plant Genetic Complementation Test Lycopersicon esculentum Phylogeny Protein Structure, Tertiary Protein Subunits Protein Transport Sequence Analysis, DNA Sequence Homology, Amino Acid Succinate-CoA Ligases Succinic Acid Cytology Enzymatic Activity Enzymes Genes Genetic Engineering Metabolism Saccharomyces Cerevisiae Tomatoes Lycopersicon esculentum Saccharomyces cerevisiae Solanum |
| description |
Despite the central importance of the TCA cycle in plant metabolism not all of the genes encoding its constituent enzymes have been functionally identified. In yeast, the heterodimeric protein succinyl CoA ligase is encoded for by two single-copy genes. Here we report the isolation of two tomato cDNAs coding for α- and one coding for the β-subunit of succinyl CoA ligase. These three cDNAs were used to complement the respective Saccharomyces cerevisiae mutants deficient in the α- and β-subunit, demonstrating that they encode functionally active polypeptides. The genes encoding for the subunits were expressed in all tissues, but most strongly in floral and leaf tissues, with equivalent expression of the two α-subunit genes being expressed to equivalent levels in all tissues. In all instances GFP fusion expression studies confirmed an expected mitochondrial location of the proteins encoded. Following the development of a novel assay to measure succinyl CoA ligase activity, in the direction of succinate formation, the evaluation of the maximal catalytic activities of the enzyme in a range of tissues revealed that these paralleled those of mRNA levels. We also utilized this assay to perform a preliminary characterisation of the regulatory properties of the enzyme suggesting allosteric control of this enzyme which may regulate flux through the TCA cycle in a manner consistent with its position therein. © Springer 2005. |
| format |
JOUR |
| author |
Studart-Guimarães, C. Gibon, Y. Frankel, N. Wood, C.C. Zanor, M.I. Fernie, A.R. Carrari, F. |
| author_facet |
Studart-Guimarães, C. Gibon, Y. Frankel, N. Wood, C.C. Zanor, M.I. Fernie, A.R. Carrari, F. |
| author_sort |
Studart-Guimarães, C. |
| title |
Identification and characterisation of the α and β subunits of succinyl CoA ligase of tomato |
| title_short |
Identification and characterisation of the α and β subunits of succinyl CoA ligase of tomato |
| title_full |
Identification and characterisation of the α and β subunits of succinyl CoA ligase of tomato |
| title_fullStr |
Identification and characterisation of the α and β subunits of succinyl CoA ligase of tomato |
| title_full_unstemmed |
Identification and characterisation of the α and β subunits of succinyl CoA ligase of tomato |
| title_sort |
identification and characterisation of the α and β subunits of succinyl coa ligase of tomato |
| url |
http://hdl.handle.net/20.500.12110/paper_01674412_v59_n5_p781_StudartGuimaraes |
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