Discovery and biochemical characterization of enzymes completing the 4-hydroxyphenylacetate pathway in Acinetobacter baumannii TH
Issued Date
2025-12-01
Resource Type
ISSN
00219258
eISSN
1083351X
Scopus ID
2-s2.0-105023108964
Pubmed ID
41203124
Journal Title
Journal of Biological Chemistry
Volume
301
Issue
12
Rights Holder(s)
SCOPUS
Bibliographic Citation
Journal of Biological Chemistry Vol.301 No.12 (2025)
Suggested Citation
Chinantuya W., Kungchuai K., Chaiyen P., Maenpuen S., Tinikul R. Discovery and biochemical characterization of enzymes completing the 4-hydroxyphenylacetate pathway in Acinetobacter baumannii TH. Journal of Biological Chemistry Vol.301 No.12 (2025). doi:10.1016/j.jbc.2025.110917 Retrieved from: https://repository.li.mahidol.ac.th/handle/123456789/113389
Title
Discovery and biochemical characterization of enzymes completing the 4-hydroxyphenylacetate pathway in Acinetobacter baumannii TH
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Corresponding Author(s)
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Abstract
A metabolic pathway for degrading 4-hydroxyphenylacetate (4-HPA) is crucial for environmental and pathogenic microbes to assimilate aromatic compounds. The 4-HPA degradation pathway in Acinetobacter baumannii TH comprises multiple reactions that are not fully understood. Enzymes involved in the first two steps (a two-component 4-HPA-3-hydroxylase, and 3,4-dihydroxyphenylacetate 2,3-dioxygenase) and the last two steps (4-hydroxy-2-keto-heptane-1,7-dioate aldolase and succinic semialdehyde (SSA) dehydrogenase (SSADH)) have been identified and studied, while the enzymes functioning in the middle of the pathway remain uncharacterized. Here, we identified products associated with individual enzymes including 5-carboxymethyl-2-hydroxymuconate-semialdehyde (CHMS) dehydrogenase (CHMSD), 5-carboxymethyl-2-hydroxymuconate (CHM) isomerase (CHMI), five-oxo-pent-3-ene-1,2,5-tricarboxylate (OPET) decarboxylase (OPETD), 2-hydroxy-hept-2,4-diene-1,7-dioate (HHDD) isomerase (HHDDI) and two-oxo-hept-3-ene-1,7-dioate (OHED) hydratase (OHEDH). We used enzymatically synthesized OPET (a tri-acid) to probe the decarboxylation step and found that the highest decarboxylation efficiency was achieved when OPETD, CHMI, and HHDDI were all present in the reaction. We demonstrated that CHMI was responsible for tri-acid tautomerization, while the protein-protein interactions between OPETD and HHDDI enhanced the decarboxylation by OPETD to generate OHED (a di-acid). OHEDH is distinct from other hydratases in that it requires Mn<sup>2+</sup> as a cofactor. Notably, besides CHMS, CHMSD can use SSA, a substrate of SSADH, suggesting that CHMSD can substitute for SSADH to generate succinate for cellular utilization. Our studies herein completely assigned the catalytic functions of all enzymes in the 4-HPA degradation pathway. The knowledge gained will be valuable for developing inhibitors targeting enzymes unique to pathogenic microbes or for constructing cascade reactions to convert lignin-derived compounds into valuable biochemicals.