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Cloning, expression and characterization of LeFRK3, the fourth tomato (Lycopersicon esculentum Mill.) gene encoding fructokinase
Year:
2004
Source of publication :
Plant Science
Authors :
Asher, Israel
;
.
Dai, Nir
;
.
German, Marcelo A.
;
.
Granot, David
;
.
Petreikov, Marina
;
.
Schaffer, Arthur
;
.
Volume :
166
Co-Authors:
German, M.A., Institute of Field and Garden Crops, Agricultural Research Organization, Volcani Center, Bet Dagan 50250, Israel
Asher, I., Institute of Field and Garden Crops, Agricultural Research Organization, Volcani Center, Bet Dagan 50250, Israel
Petreikov, M., Institute of Field and Garden Crops, Agricultural Research Organization, Volcani Center, Bet Dagan 50250, Israel
Dai, N., Institute of Field and Garden Crops, Agricultural Research Organization, Volcani Center, Bet Dagan 50250, Israel
Schaffer, A.A., Institute of Field and Garden Crops, Agricultural Research Organization, Volcani Center, Bet Dagan 50250, Israel
Granot, D., Institute of Field and Garden Crops, Agricultural Research Organization, Volcani Center, Bet Dagan 50250, Israel
Facilitators :
From page:
285
To page:
291
(
Total pages:
7
)
Abstract:
A full-length cDNA encoding a novel fourth fructokinase, LeFRK3, was cloned from green tomato (Lycopersicon esculentum Mill.) fruits. The putative protein shares 70, 65.5 and 69% amino acid homology with the three previously identified tomato fructokinases encoded by LeFRK1, LeFRK2 and LeFRK4, respectively. This fourth fructokinase has signature patterns of the pfkB family of carbohydrate kinases as well as substrate recognition sites and an ATP-binding domain. Confirmation for its fructokinase activity was obtained by complementation of triple mutant yeast cells that are unable to phosphorylate or grow on either glucose or fructose as LeFRK3 complemented growth on fructose but not on glucose. Moreover, soluble crude protein extracts prepared from the transformed yeast cells revealed fructose but not glucose phosphorylation activity. In contrast to the LeFRK1 gene product which is inhibited neither by fructose nor by Mg, and to LeFRK2 gene product which is inhibited by both fructose and Mg, the LeFRK3 product is inhibited by fructose but not by Mg. Separation by HPLC-ion exchange chromatography pointed to the gene product of LeFRK3 as the protein responsible for the third peak of fructokinase activity (FKIII), sharing the same pattern of fructose inhibition previously identified with FKIII in tomato fruits. Mapping of tomato fructokinases indicated that LeFRK3 is located on chromosome 2, unlike LeFRK1 (chromosome 3), LeFRK2 (chromosome 6), and LeFRK4 (chromosome 10). The relative expression levels of the four known FRK genes in different tomato organs were analyzed by quantitative RT-PCR. LeFRK2 and LeFRK3 are the predominant genes expressed in all organs with LeFRK3 having the highest level of expression in leaves and apices. LeFRK4 is expressed only in stamens. This differential expression patterns combined with the different biochemical characteristics of the four FRK isozymes suggest that each plays a different role in plant development. © 2003 Elsevier Ireland Ltd. All rights reserved.
Note:
Related Files :
Amino Acids
carbohydrates
cloning
DNA
enzyme inhibition
Fruits
gene transfer
Hexose phosphorylation
Yeast
Show More
Related Content
More details
DOI :
10.1016/j.plantsci.2003.09.017
Article number:
Affiliations:
Database:
Scopus
Publication Type:
article
;
.
Language:
English
Editors' remarks:
ID:
28763
Last updated date:
02/03/2022 17:27
Creation date:
17/04/2018 00:41
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Scientific Publication
Cloning, expression and characterization of LeFRK3, the fourth tomato (Lycopersicon esculentum Mill.) gene encoding fructokinase
166
German, M.A., Institute of Field and Garden Crops, Agricultural Research Organization, Volcani Center, Bet Dagan 50250, Israel
Asher, I., Institute of Field and Garden Crops, Agricultural Research Organization, Volcani Center, Bet Dagan 50250, Israel
Petreikov, M., Institute of Field and Garden Crops, Agricultural Research Organization, Volcani Center, Bet Dagan 50250, Israel
Dai, N., Institute of Field and Garden Crops, Agricultural Research Organization, Volcani Center, Bet Dagan 50250, Israel
Schaffer, A.A., Institute of Field and Garden Crops, Agricultural Research Organization, Volcani Center, Bet Dagan 50250, Israel
Granot, D., Institute of Field and Garden Crops, Agricultural Research Organization, Volcani Center, Bet Dagan 50250, Israel
Cloning, expression and characterization of LeFRK3, the fourth tomato (Lycopersicon esculentum Mill.) gene encoding fructokinase
A full-length cDNA encoding a novel fourth fructokinase, LeFRK3, was cloned from green tomato (Lycopersicon esculentum Mill.) fruits. The putative protein shares 70, 65.5 and 69% amino acid homology with the three previously identified tomato fructokinases encoded by LeFRK1, LeFRK2 and LeFRK4, respectively. This fourth fructokinase has signature patterns of the pfkB family of carbohydrate kinases as well as substrate recognition sites and an ATP-binding domain. Confirmation for its fructokinase activity was obtained by complementation of triple mutant yeast cells that are unable to phosphorylate or grow on either glucose or fructose as LeFRK3 complemented growth on fructose but not on glucose. Moreover, soluble crude protein extracts prepared from the transformed yeast cells revealed fructose but not glucose phosphorylation activity. In contrast to the LeFRK1 gene product which is inhibited neither by fructose nor by Mg, and to LeFRK2 gene product which is inhibited by both fructose and Mg, the LeFRK3 product is inhibited by fructose but not by Mg. Separation by HPLC-ion exchange chromatography pointed to the gene product of LeFRK3 as the protein responsible for the third peak of fructokinase activity (FKIII), sharing the same pattern of fructose inhibition previously identified with FKIII in tomato fruits. Mapping of tomato fructokinases indicated that LeFRK3 is located on chromosome 2, unlike LeFRK1 (chromosome 3), LeFRK2 (chromosome 6), and LeFRK4 (chromosome 10). The relative expression levels of the four known FRK genes in different tomato organs were analyzed by quantitative RT-PCR. LeFRK2 and LeFRK3 are the predominant genes expressed in all organs with LeFRK3 having the highest level of expression in leaves and apices. LeFRK4 is expressed only in stamens. This differential expression patterns combined with the different biochemical characteristics of the four FRK isozymes suggest that each plays a different role in plant development. © 2003 Elsevier Ireland Ltd. All rights reserved.
Scientific Publication
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