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A dual positional specific lipoxygenase functions in the generation of flavor compounds during climacteric ripening of apple.

Schiller D, Contreras C, Vogt J, Dunemann F, Defilippi BG, Beaudry R, Schwab W - Hortic Res (2015)

Bottom Line: Site-directed mutagenesis of Gly567 to an alanine converted the dual positional specific LOX1:Md:1a to an enzyme with a high specificity for 9(S)-hydroperoxide formation.The high expression level of the corresponding MdLOX1a gene in stored apple fruit, the genetic association with a quantitative trait locus for fruit ester and the remarkable agreement in regio- and stereoselectivity of the LOX1:Md:1a reaction with the overall LOX activity found in mature apple fruits, suggest a major physiological function of LOX1:Md:1a during climacteric ripening of apples.While LOX1:Md:1c, LOX2:Md:2a and LOX2:Md:2b may contribute to aldehyde production in immature fruit upon cell disruption our results furnish additional evidence that LOX1:Md:1a probably regulates the availability of precursors for ester production in intact fruit tissue.

View Article: PubMed Central - PubMed

Affiliation: Biotechnology of Natural Products, Technische Universität München , Liesel-Beckmann-Str. 1, D-85354 Freising, Germany.

ABSTRACT
Lipoxygenase (LOX) is an important contributor to the formation of aroma-active C6 aldehydes in apple (Malus × domestica) fruit upon tissue disruption but little is known about its role in autonomously produced aroma volatiles from intact tissue. We explored the expression of 22 putative LOX genes in apple throughout ripening, but only six LOXs were expressed in a ripening-dependent manner. Recombinant LOX1:Md:1a, LOX1:Md:1c, LOX2:Md:2a and LOX2:Md:2b proteins showed 13/9-LOX, 9-LOX, 13/9-LOX and 13-LOX activity with linoleic acid, respectively. While products of LOX1:Md:1c and LOX2:Md:2b were S-configured, LOX1:Md:1a and LOX2:Md:2a formed 13(R)-hydroperoxides as major products. Site-directed mutagenesis of Gly567 to an alanine converted the dual positional specific LOX1:Md:1a to an enzyme with a high specificity for 9(S)-hydroperoxide formation. The high expression level of the corresponding MdLOX1a gene in stored apple fruit, the genetic association with a quantitative trait locus for fruit ester and the remarkable agreement in regio- and stereoselectivity of the LOX1:Md:1a reaction with the overall LOX activity found in mature apple fruits, suggest a major physiological function of LOX1:Md:1a during climacteric ripening of apples. While LOX1:Md:1c, LOX2:Md:2a and LOX2:Md:2b may contribute to aldehyde production in immature fruit upon cell disruption our results furnish additional evidence that LOX1:Md:1a probably regulates the availability of precursors for ester production in intact fruit tissue.

No MeSH data available.


Related in: MedlinePlus

Expression of selected LOX genes. Relative expression levels of putative 9-LOX (upper panel) and 13-LOX (lower panel) transcripts using qPCR for peel tissue from ‘Jonagold’ apple fruit during ripening. Expression is normalized relative to the earliest (preclimacteric) developmental stage evaluated. Time points selected are the same as those depicted in Figure 2, with the onset of autocatalytic ethylene production occurring between stages 4 and 5.
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fig3: Expression of selected LOX genes. Relative expression levels of putative 9-LOX (upper panel) and 13-LOX (lower panel) transcripts using qPCR for peel tissue from ‘Jonagold’ apple fruit during ripening. Expression is normalized relative to the earliest (preclimacteric) developmental stage evaluated. Time points selected are the same as those depicted in Figure 2, with the onset of autocatalytic ethylene production occurring between stages 4 and 5.

Mentions: qPCR was performed on the 6 LOX genes that showed ripening-dependent patterns, and additionally included MdLOX2a due to its lack of variation during ripening to provide additional support for the accuracy of the RT-PCR results. In each instance, the RT-PCR and qPCR results yielded similar results (Figure 3). In the putative 13-LOX group, MdLOX2a exhibited little change or a slight decrease in expression during ripening, again MdLOX4a evidenced a sharp increase at stage 5, and MdLOX6a and MdLOX6b were downregulated during ripening. Within the putative 9-LOX group, MdLOX7c was downregulated, and MdLOX7a and MdLOX1a were upregulated as ripening progressed. MdLOX1a expression increased more than 100-fold throughout the course of ripening, with the initial increase occurring between developmental stages 3 and 4, concomitant with the first detectable emissions of hexyl esters.


A dual positional specific lipoxygenase functions in the generation of flavor compounds during climacteric ripening of apple.

Schiller D, Contreras C, Vogt J, Dunemann F, Defilippi BG, Beaudry R, Schwab W - Hortic Res (2015)

Expression of selected LOX genes. Relative expression levels of putative 9-LOX (upper panel) and 13-LOX (lower panel) transcripts using qPCR for peel tissue from ‘Jonagold’ apple fruit during ripening. Expression is normalized relative to the earliest (preclimacteric) developmental stage evaluated. Time points selected are the same as those depicted in Figure 2, with the onset of autocatalytic ethylene production occurring between stages 4 and 5.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4595979&req=5

fig3: Expression of selected LOX genes. Relative expression levels of putative 9-LOX (upper panel) and 13-LOX (lower panel) transcripts using qPCR for peel tissue from ‘Jonagold’ apple fruit during ripening. Expression is normalized relative to the earliest (preclimacteric) developmental stage evaluated. Time points selected are the same as those depicted in Figure 2, with the onset of autocatalytic ethylene production occurring between stages 4 and 5.
Mentions: qPCR was performed on the 6 LOX genes that showed ripening-dependent patterns, and additionally included MdLOX2a due to its lack of variation during ripening to provide additional support for the accuracy of the RT-PCR results. In each instance, the RT-PCR and qPCR results yielded similar results (Figure 3). In the putative 13-LOX group, MdLOX2a exhibited little change or a slight decrease in expression during ripening, again MdLOX4a evidenced a sharp increase at stage 5, and MdLOX6a and MdLOX6b were downregulated during ripening. Within the putative 9-LOX group, MdLOX7c was downregulated, and MdLOX7a and MdLOX1a were upregulated as ripening progressed. MdLOX1a expression increased more than 100-fold throughout the course of ripening, with the initial increase occurring between developmental stages 3 and 4, concomitant with the first detectable emissions of hexyl esters.

Bottom Line: Site-directed mutagenesis of Gly567 to an alanine converted the dual positional specific LOX1:Md:1a to an enzyme with a high specificity for 9(S)-hydroperoxide formation.The high expression level of the corresponding MdLOX1a gene in stored apple fruit, the genetic association with a quantitative trait locus for fruit ester and the remarkable agreement in regio- and stereoselectivity of the LOX1:Md:1a reaction with the overall LOX activity found in mature apple fruits, suggest a major physiological function of LOX1:Md:1a during climacteric ripening of apples.While LOX1:Md:1c, LOX2:Md:2a and LOX2:Md:2b may contribute to aldehyde production in immature fruit upon cell disruption our results furnish additional evidence that LOX1:Md:1a probably regulates the availability of precursors for ester production in intact fruit tissue.

View Article: PubMed Central - PubMed

Affiliation: Biotechnology of Natural Products, Technische Universität München , Liesel-Beckmann-Str. 1, D-85354 Freising, Germany.

ABSTRACT
Lipoxygenase (LOX) is an important contributor to the formation of aroma-active C6 aldehydes in apple (Malus × domestica) fruit upon tissue disruption but little is known about its role in autonomously produced aroma volatiles from intact tissue. We explored the expression of 22 putative LOX genes in apple throughout ripening, but only six LOXs were expressed in a ripening-dependent manner. Recombinant LOX1:Md:1a, LOX1:Md:1c, LOX2:Md:2a and LOX2:Md:2b proteins showed 13/9-LOX, 9-LOX, 13/9-LOX and 13-LOX activity with linoleic acid, respectively. While products of LOX1:Md:1c and LOX2:Md:2b were S-configured, LOX1:Md:1a and LOX2:Md:2a formed 13(R)-hydroperoxides as major products. Site-directed mutagenesis of Gly567 to an alanine converted the dual positional specific LOX1:Md:1a to an enzyme with a high specificity for 9(S)-hydroperoxide formation. The high expression level of the corresponding MdLOX1a gene in stored apple fruit, the genetic association with a quantitative trait locus for fruit ester and the remarkable agreement in regio- and stereoselectivity of the LOX1:Md:1a reaction with the overall LOX activity found in mature apple fruits, suggest a major physiological function of LOX1:Md:1a during climacteric ripening of apples. While LOX1:Md:1c, LOX2:Md:2a and LOX2:Md:2b may contribute to aldehyde production in immature fruit upon cell disruption our results furnish additional evidence that LOX1:Md:1a probably regulates the availability of precursors for ester production in intact fruit tissue.

No MeSH data available.


Related in: MedlinePlus