Structure–function analyses of two plant meso-diaminopimelate decarboxylase isoforms reveal that active-site gating provides stereochemical control
| Autor: | Mary M. Leeman, Michael R. Oliver, Rachel A. North, Austin J. Bartl, Müge Kasanmascheff, Renwick C. J. Dobson, Hironori Suzuki, Michael D. W. Griffin, Katherine A. Donovan, André O. Hudson, Jennifer M. Crowther, Penelope J. Cross |
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| Rok vydání: | 2019 |
| Předmět: |
0301 basic medicine
Gene isoform DAPDC Arabidopsis thaliana Carboxy-Lyases Decarboxylation small-angle X-ray scattering (SAXS) Arabidopsis Crystallography X-Ray Biochemistry Diaminopimelate decarboxylase 03 medical and health sciences Protein Domains Catalytic Domain meso-diaminopimelate decarboxylase Lysine biosynthesis Molecular Biology pyridoxal phosphate X-ray crystallography chemistry.chemical_classification 030102 biochemistry & molecular biology biology Arabidopsis Proteins Chemistry fungi stereochemistry Active site Cell Biology biology.organism_classification Enzyme structure Complementation 030104 developmental biology Enzyme Enzymology biology.protein analytical ultracentrifugation (AUC) |
| Zdroj: | Crowther, J M, Cross, P J, Oliver, M R, Leeman, M M, Bartl, A J, Weatherhead, A W, North, R A, Donovan, K A, Griffin, M D W, Suzuki, H, Hudson, A O, Kasanmascheff, M & Dobson, R C J 2019, ' Structure–function analyses of two plant meso-diaminopimelate decarboxylase isoforms reveal that active-site gating provides stereochemical control ', Journal of Biological Chemistry, vol. 294, no. 21, pp. 8505-8515 . https://doi.org/10.1074/jbc.RA118.006825 J Biol Chem |
| ISSN: | 0021-9258 |
| Popis: | meso-Diaminopimelate decarboxylase catalyzes the decarboxylation of meso-diaminopimelate, the final reaction in the diaminopimelate l-lysine biosynthetic pathway. It is the only known pyridoxal-5-phosphate–dependent decarboxylase that catalyzes the removal of a carboxyl group from a d-stereocenter. Currently, only prokaryotic orthologs have been kinetically and structurally characterized. Here, using complementation and kinetic analyses of enzymes recombinantly expressed in Escherichia coli, we have functionally tested two putative eukaryotic meso-diaminopimelate decarboxylase isoforms from the plant species Arabidopsis thaliana. We confirm they are both functional meso-diaminopimelate decarboxylases, although with lower activities than those previously reported for bacterial orthologs. We also report in-depth X-ray crystallographic structural analyses of each isoform at 1.9 and 2.4 Å resolution. We have captured the enzyme structure of one isoform in an asymmetric configuration, with one ligand-bound monomer and the other in an apo-form. Analytical ultracentrifugation and small-angle X-ray scattering solution studies reveal that A. thaliana meso-diaminopimelate decarboxylase adopts a homodimeric assembly. On the basis of our structural analyses, we suggest a mechanism whereby molecular interactions within the active site transduce conformational changes to the active-site loop. These conformational differences are likely to influence catalytic activity in a way that could allow for d-stereocenter selectivity of the substrate meso-diaminopimelate to facilitate the synthesis of l-lysine. In summary, the A. thaliana gene loci At3g14390 and At5g11880 encode functional. meso-diaminopimelate decarboxylase enzymes whose structures provide clues to the stereochemical control of the decarboxylation reaction catalyzed by these eukaryotic proteins |
| Databáze: | OpenAIRE |
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