Benzoate-CoA ligase contributes to the biosynthesis of biphenyl phytoalexins in elicitor-treated pear cell cultures
Benzoate-Coenzyme A ligase enzyme activity catalyzing the conversion of free benzoic acid to benzoyl-CoA was detected and biochemically characterized in the elicitor-treated pear cell cultures.
Asian pear (Pyrus pyrifolia) is an economically and nutritionally important fruit-bearing tree of the subtribe Malinae. Upon pathogen attack, pears produce unique benzoate-derived biphenyl phytoalexins. The upstream biosynthesis of the biphenyl in Malinae is still incomplete. Previously, protein preparations from yeast extract-treated pear cultures were able to convert l-phenylalanine to cinnamic acid catalyzed by the activity of the phenylalanine ammonia lyase. The same extract was able to perform a C2 side-chain cleavage of cinnamic acid to benzaldehyde followed by oxidation of the latter to benzoic acid owing to the molecularly-undefined benzaldehyde synthase and benzaldehyde dehydrogenase activities, respectively. The biosynthesis of biphenyls starts with benzoate-Coenzyme A ligase (BZL), which converts benzoic acid to benzoyl-CoA. Subsequently, the previously-defined biphenyl synthase uses benzoyl-CoA to form the biphenyls. The current study reports the first time detection and characterization of BZL activity in elicitor-treated pear cell cultures. The preferred substrate was benzoic acid (Km = 62 ± 4 µM). Magnesium or manganese was prerequisite for the activity, which was enhanced by ~ 70% in the presence of potassium. Maximum BZL activity was observed 18 h post elicitation, which is in agreement with the coordinate induction reported for the enzymes in the same pathway. The induced BZL activity preceded the accumulation of biphenyls supporting its involvement in their biosynthesis.
KeywordsBenzoate-coenzyme A ligase Benzoic acid Biphenyl Cinnamate-CoA ligase Pyrus pyrifolia
This research work was supported by a start up research grant (FIG 100624 to D. Sircar) from the Indian Institute of Technology Roorkee. SSS is thankful to MHRD-research assistantship (MHRD02-23-200-429) from Indian Institute of Technology Roorkee for perusing his doctoral studies. We thank Dr. Till Beuerle (Technische Universität Braunschweig, Germany) for ESI–MS analyses.
Author contribution statement
DS conceived and designed the study. SSS performed experiments, data processing and analyses. MG and DS performed data analyses and wrote the manuscript.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflicts of interest.
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