1QMG

Acetohydroxyacid isomeroreductase complexed with its reaction product dihydroxy-methylvalerate, manganese and ADP-ribose.

Summary for 1QMG

• Entry DOI: 10.2210/pdb1qmg/pdb
• Related: 1YVE
• Descriptor: ACETOHYDROXY-ACID ISOMEROREDUCTASE, 2'-MONOPHOSPHOADENOSINE-5'-DIPHOSPHORIBOSE, MANGANESE (II) ION, ... (6 entities in total)
• Functional Keywords: oxidoreductase, branched chain amino acid biosynthesis, reaction product, manganese, adp-ribose
• Biological source: SPINACIA OLERACEA (SPINACH)
• Cellular location: Plastid, chloroplast: Q01292
• Total number of polymer chains: 4
• Total formula weight: 231887.71

Authors

Thomazeau, K.,Dumas, R.,Halgand, F.,Douce, R.,Biou, V. (deposition date: 1999-09-28, release date: 2000-03-31, Last modification date: 2023-12-13)

Primary citation

Thomazeau, K.,Dumas, R.,Halgand, F.,Forest, E.,Douce, R.,Biou, V.
Structure of Spinach Acetohydroxyacid Isomeroreductase Complexed with its Product of Reaction Dihydroxy-Methylvalerate, Manganese and Adp-Ribose
Acta Crystallogr.,Sect.D, 56:389-, 2000

PubMed Abstract: Acetohydroxyacid isomeroreductase catalyses a two-step reaction composed of an alkyl migration followed by an NADPH-dependent reduction. Both steps require a divalent cation and the first step has a strong preference for magnesium. Manganese ions are highly unfavourable to the reaction: only 3% residual activity is observed in the presence of this cation. Acetohydroxyacid isomeroreductase has been crystallized with its substrate, 2-aceto-2-hydroxybutyrate (AHB), Mn(2+) and NADPH. The 1.6 A resolution electron-density map showed the reaction product (2,3-dihydroxy-3-methylvalerate, DHMV) and a density corresponding to (phospho)-ADP-ribose instead of the whole NADP(+). This is one of the few structures of an enzyme complexed with its reaction product. The structure of this complex was refined to an R factor of 19.3% and an R(free) of 22.5%. The overall structure of the enzyme is very similar to that of the complex with the reaction-intermediate analogue IpOHA [N-hydroxy-N-isopropyloxamate; Biou et al. (1997), EMBO J. 16, 3405-3415]. However, the active site shows some differences: the nicotinamide is cleaved and the surrounding amino acids have rearranged accordingly. Comparison between the structures corresponding to the reaction intermediate and to the end of the reaction allowed the proposal of a reaction scheme. Taking this result into account, the enzyme was crystallized with Ni(2+) and Zn(2+), for which only 0.02% residual activity were measured; however, the crystals of AHB/Zn/NADPH and of AHB/Ni/NADPH also contain the reaction product. Moreover, mass-spectrometry measurements confirmed the -cleavage of nicotinamide.

PubMed: 10739911
DOI: 10.1107/S0907444900001694

Experimental method

X-RAY DIFFRACTION (1.6 Å)