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Identification, cloning and expression analysis of strawberry ( Fragaria × ananassa ) mitochondrial citrate synthase and mitochondrial malate dehydrogenase
Author(s) -
Iannetta Pietro P. M.,
Escobar Nieves Medina,
Ross Heather A.,
Souleyre Edwige J. F.,
Hancock Robert D.,
Witte ClausPeter,
Davies Howard V.
Publication year - 2004
Publication title -
physiologia plantarum
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.351
H-Index - 146
eISSN - 1399-3054
pISSN - 0031-9317
DOI - 10.1111/j.0031-9317.2004.00302.x
Subject(s) - malate dehydrogenase , biochemistry , citrate synthase , biology , fragaria , enzyme , atp citrate lyase , dehydrogenase , ripening , sucrose phosphate synthase , microbiology and biotechnology , sucrose synthase , botany , invertase
Salt‐extractable proteins from the cell walls of immature and ripe strawberry ( Fragaria × ananassa Duch. cv. Elsanta) fruit were separated using two‐dimensional polyacrylamide gel electrophoresis. Seven polypeptides (enzymes) were characterized from their N‐terminal sequences: (1) glyceraldhyde‐3‐phosphate dehydrogenase (EC 1.2.1.12); (2) triose phosphate isomerase (TPI; EC 5.3.1.1); (3) mitochondrial malate dehydrogenase (mMDH; EC 1.1.1.37); (4) NADH glutamate dehydrogenase (EC 1.4.1.3); (5) chalcone synthase (ChS; EC 2.3.1.74); (6) mitochondrial citrate synthase (mCS; EC 4.1.3.7); and (7) UDP glucose:flavonoid 3‐ O ‐glucosyltransferase (UDPG:FGT; EC 2.4.1.91). The sequenced polypeptides identified only cytosolic proteins, two of which (ChS and UDPG:FGT) had already been identified as being up‐regulated in ripening (strawberry) fruit and important contributors to ripe fruit character. Our focus was therefore diverted to the enzymes mMDH and mCS for further molecular characterization as potentially important determinants of fruit flavour via regulation of the sugar : acid balance. Citrate synthase (CS) and malate dehydrogenase (MDH) enzyme activities increased substantially during ripening, as did citrate and malate contents. The increase in CS activity is supported by western blot analysis. One strawberry mCS ( Fa‐mCS‐I ) and two mMDH ( Fa‐mMDH‐I and ‐II ) cDNAs were cloned that were 77, 82 and 53% identical (respectively) to sequences from other plant sources. Northern analysis showed that CS and MDH expression did not correlate with enzyme activities and these findings are discussed.