TY - JOUR
T1 - Discovery of Novel p-Hydroxybenzoate- m-hydroxylase, Protocatechuate 3,4 Ring-Cleavage Dioxygenase, and Hydroxyquinol 1,2 Ring-Cleavage Dioxygenase from the Filamentous Fungus Aspergillus niger
AU - Lubbers, Ronnie J.M.
AU - Dilokpimol, Adiphol
AU - Peng, Mao
AU - Visser, Jaap
AU - Mäkelä, Miia R.
AU - Hildén, Kristiina S.
AU - De Vries, Ronald P.
N1 - Funding Information:
R.J.M.L. conducted the experiments, analyzed the data, and wrote the manuscript. M.P. processed and analyzed RNA sequencing data. R.P.d.V. conceived and supervised the overall project. All authors commented on the manuscript. This project was supported through FALCON by the European Union’s Horizon 2020 research and innovation programme under grant agreement no. 720918. The authors declare no competing financial interest.
Publisher Copyright:
Copyright © 2019 American Chemical Society.
PY - 2019/12/2
Y1 - 2019/12/2
N2 - The aromatic compound benzoic acid and its derivatives are frequently used preservatives in food and beverages due to their strong antimicrobial properties. Benzoic acid naturally occurs in plants and serves as a building block for primary and secondary metabolites. Several Aspergilli are able to degrade benzoic acid to p-hydroxybenzoic acid by p-hydroxylation followed by m-hydroxylation to protocatechuic acid. The aromatic ring of protocatechuic acid is then cleaved and further converted through the oxoadipate pathway where it is used as a carbon source. The benzoic acid pathway is well studied in the filamentous fungus Aspergillus niger; however, only one enzyme, benzoate-4-monooxygenase, has been characterized. In this study, three novel enzymes of the benzoic acid pathway were identified and characterized. Transcriptome analysis of A. niger on p-hydroxybenzoic acid, protocatechuic acid, and the related compounds p-coumaric acid and caffeic acid revealed two genes of unknown function, which were highly expressed on all four aromatic compounds. Analysis of A. niger deletion mutants of these genes revealed their essential role in the utilization of benzoic acid, p-hydroxybenzoic acid, and protocatechuic acid. Biochemical analysis confirmed that the corresponding enzymes function as the p-hydroxybenzoate-m-hydroxylase and protocatechuate 3,4 ring-cleavage dioxygenase in the benzoic acid pathway. In addition, a hydroxyquinol 1,2 ring-cleavage dioxygenase, involved in an alternative protocatechuic acid metabolic pathway, was identified.
AB - The aromatic compound benzoic acid and its derivatives are frequently used preservatives in food and beverages due to their strong antimicrobial properties. Benzoic acid naturally occurs in plants and serves as a building block for primary and secondary metabolites. Several Aspergilli are able to degrade benzoic acid to p-hydroxybenzoic acid by p-hydroxylation followed by m-hydroxylation to protocatechuic acid. The aromatic ring of protocatechuic acid is then cleaved and further converted through the oxoadipate pathway where it is used as a carbon source. The benzoic acid pathway is well studied in the filamentous fungus Aspergillus niger; however, only one enzyme, benzoate-4-monooxygenase, has been characterized. In this study, three novel enzymes of the benzoic acid pathway were identified and characterized. Transcriptome analysis of A. niger on p-hydroxybenzoic acid, protocatechuic acid, and the related compounds p-coumaric acid and caffeic acid revealed two genes of unknown function, which were highly expressed on all four aromatic compounds. Analysis of A. niger deletion mutants of these genes revealed their essential role in the utilization of benzoic acid, p-hydroxybenzoic acid, and protocatechuic acid. Biochemical analysis confirmed that the corresponding enzymes function as the p-hydroxybenzoate-m-hydroxylase and protocatechuate 3,4 ring-cleavage dioxygenase in the benzoic acid pathway. In addition, a hydroxyquinol 1,2 ring-cleavage dioxygenase, involved in an alternative protocatechuic acid metabolic pathway, was identified.
KW - benzoic acid pathway
KW - flavoprotein
KW - intradiol ring fission
UR - http://www.scopus.com/inward/record.url?scp=85075159020&partnerID=8YFLogxK
U2 - 10.1021/acssuschemeng.9b04918
DO - 10.1021/acssuschemeng.9b04918
M3 - Article
AN - SCOPUS:85075159020
SN - 2168-0485
VL - 7
SP - 19081
EP - 19089
JO - ACS Sustainable Chemistry and Engineering
JF - ACS Sustainable Chemistry and Engineering
IS - 23
ER -