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Uncovering the functional residues of Arabidopsis isoprenoid biosynthesis enzyme HDS.

ABSTRACT: The methylerythritol phosphate (MEP) pathway is responsible for producing isoprenoids, metabolites with essential functions in the bacterial kingdom and plastid-bearing organisms including plants and Apicomplexa. Additionally, the MEP-pathway intermediate methylerythritol cyclodiphosphate (MEcPP) serves as a plastid-to-nucleus retrograde signal. A suppressor screen of the high MEcPP accumulating mutant plant (ceh1) led to the isolation of 3 revertants (designated Rceh1-3) resulting from independent intragenic substitutions of conserved amino acids in the penultimate MEP-pathway enzyme, hydroxymethylbutenyl diphosphate synthase (HDS). The revertants accumulate varying MEcPP levels, lower than that of ceh1, and exhibit partial or full recovery of MEcPP-mediated phenotypes, including stunted growth and induced expression of stress response genes and the corresponding metabolites. Structural modeling of HDS and ligand docking spatially position the substituted residues at the MEcPP binding pocket and cofactor binding domain of the enzyme. Complementation assays confirm the role of these residues in suppressing the ceh1 mutant phenotypes, albeit to different degrees. In vitro enzyme assays of wild type and HDS variants exhibit differential activities and reveal an unanticipated mismatch between enzyme kinetics and the in vivo MEcPP levels in the corresponding Rceh lines. Additional analyses attribute the mismatch, in part, to the abundance of the first and rate-limiting MEP-pathway enzyme, DXS, and further suggest MEcPP as a rheostat for abundance of the upstream enzyme instrumental in fine-tuning of the pathway flux. Collectively, this study identifies critical residues of a key MEP-pathway enzyme, HDS, valuable for synthetic engineering of isoprenoids, and as potential targets for rational design of antiinfective drugs.

SUBMITTER: Wang JZ

PROVIDER: S-EPMC6955319 | biostudies-literature | 2020 Jan

REPOSITORIES: biostudies-literature

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Uncovering the functional residues of Arabidopsis isoprenoid biosynthesis enzyme HDS.

Wang Jin-Zheng JZ Lei Yongxing Y Xiao Yanmei Y He Xiang X Liang Jiubo J Jiang Jishan J Dong Shangzhi S Ke Haiyan H Leon Patricia P Zerbe Philipp P Xiao Youli Y Dehesh Katayoon K

Proceedings of the National Academy of Sciences of the United States of America 20191226 1

The methylerythritol phosphate (MEP) pathway is responsible for producing isoprenoids, metabolites with essential functions in the bacterial kingdom and plastid-bearing organisms including plants and Apicomplexa. Additionally, the MEP-pathway intermediate methylerythritol cyclodiphosphate (MEcPP) serves as a plastid-to-nucleus retrograde signal. A suppressor screen of the high MEcPP accumulating mutant plant (ceh1) led to the isolation of 3 revertants (designated Rceh1-3) re ...[more]

PMID: 31879352

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Dataset Information

Uncovering the functional residues of Arabidopsis isoprenoid biosynthesis enzyme HDS.

Publications

Uncovering the functional residues of <i>Arabidopsis</i> isoprenoid biosynthesis enzyme HDS.

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