Cloning and characterization of a cDNA encoding hexokinase from tomato
| Autor: | Thierry Menu, Christophe Rothan, C. Etienne, Agnès Destrac-Irvine, Bérénice Ricard, Marina Petreikov, Nir Dai, Arthur A. Schaffer, David Granot |
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| Přispěvatelé: | Station de physiologie végétale, Institut National de la Recherche Agronomique (INRA), Institute of Field and Garden Crops, Agricultural Research Organization, Agr Res Org Volcani Ctr |
| Jazyk: | angličtina |
| Rok vydání: | 2001 |
| Předmět: |
0106 biological sciences
[SDV]Life Sciences [q-bio] Saccharomyces cerevisiae RT-PCR Plant Science 01 natural sciences Lycopersicon [SDV.GEN.GPL]Life Sciences [q-bio]/Genetics/Plants genetics 03 medical and health sciences chemistry.chemical_compound ADNC Complementary DNA Genetics 030304 developmental biology 0303 health sciences Hexokinase biology cDNA library food and beverages Fructose General Medicine BIOLOGIE MOLECULAIRE biology.organism_classification Yeast Open reading frame Biochemistry chemistry METABOLISME DES GLUCIDES Sugar phosphory Agronomy and Crop Science cDNA 010606 plant biology & botany lation Lycopersicon esculentum |
| Zdroj: | Plant Science Plant Science, Elsevier, 2001, pp.209-218 Plant Science, Elsevier, 2001, 160 (2), pp.209-218. ⟨10.1016/S0168-9452(00)00332-0⟩ |
| ISSN: | 0168-9452 |
| DOI: | 10.1016/S0168-9452(00)00332-0⟩ |
| Popis: | International audience; Two different partial sequences encoding putative hexokinase (HXK, ATP: hexose-6-phosphotransferase: EC 2.7.1.1) were isolated from tomato (Lycopersicon esculentum) by RT-PCR using degenerate primers. Southern blot analysis suggested the existence of two divergent HXK genes. A complete cDNA of one HXK was isolated by screening a cDNA library prepared from young cherry tomato fruit. The 1770 bp cDNA of LeHXK2 contained an open reading frame encoding a 496 amino acid protein that has 69% identity with the two Arabidopsis HXKs, 83 and 85% identity with potato StHXK1 and tobacco NtHXK, respectively. However, this clone had 97% amino acid identity with potato StHXK2 and, therefore, was named LeHXK2. LeHXK2 cDNA was expressed in a triple mutant yeast (Saccharomyces cerevisiae) strain which lacked the ability to phosphorylate glucose and fructose and, therefore, was unable to grow on these sugars as carbon sources. Mutant cells expressing LeHXK2 grew on both glucose and fructose with shorter doubling time on glucose. The kinetic properties of LeHXK2 expressed in yeast were determined after the purification of LeHXK2 by HPLC-ion exchange chromatography, confirming the identity of LeHXK2 as hexokinase with higher affinity to glucose. LeHXK2 mRNA was detected by RT-PCR expression analysis in all organs and tissues and at all stages of fruit development. However, semi-quantitative RT-PCR analysis showed that LeHXK2 was most highly expressed in flowers. |
| Databáze: | OpenAIRE |
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