Structure, Catalysis and Supramolecular Assembly of Adenylate Kinase from Maize

Abstract

The crystal structure of adenylate kinase from maize ligated with an inhibitor has been determined by molecular replacement and refined to 3.5-Å resolution. The enzyme keeps the ATP/ADP/AMP equilibrium in the cell. In the C 4 plant maize, it has the special task to recycle the AMP produced in large amounts in primary CO 2 assimilation. The established structure explains the side reaction with CMP. Moreover, it shows infinite rods that can be readily discerned in the crystal packing. In comparison with homologues, two structural differences that are crucial for this supramolecular assembly are evident. We propose that the rods represent a natural inactive storage form that assembles at night when maize stops CO 2 assimilation and thus most of the AMP production in its C 4 cycle. The enzyme is particularly abundant in mesophyll chloroplasts, where such an assembly would release appreciable amounts of water that can be used in other processes during the night.Keywords : C 4 plant metabolism; crystal contact; dual substrate specificity; supramolecular structure ; X-ray structure analysis.Adenylate kinases are ubiquitous monomeric enzymes that tity) to the mammalian isoenzyme-2 from the mitochondrial intermembrane space. catalyze the reaction AKM has been isolated from leaves [3, 4, 10] as a homogeMg 2ϩ ATP ϩ AMP < Mg 2ϩ ADP ϩ ADP.neous protein containing 222 residues with a molecular mass of 24 867 Da [9]. Besides AMP, the enzyme accepts also CMP at They belong to the well-characterized family of nucleosidemonophosphate kinases [1,2]. Maize (Zea mays) is a C 4 plant a relative activity of 10% [4]. Here, we report the crystal structure of AKM in complex with the inhibitor [11] adenosineand contains a particularly large amount of adenylate kinase (AKM), i.e. about 50-times more than C 3 plants [3Ϫ5]. Like the (5′)pentaphospho(5′)adenosine (Ap 5 A), explain the CMP activity, and deduce a putative natural supramolecular assembly. other C 4 plants, maize needs a high catalytic efficiency because its CO 2 assimilation produces stoichiometric amounts of AMP in the mesophyll chloroplasts, which have to be recycled quickly EXPERIMENTAL PROCEDURES [5Ϫ7]. The respective reactions are:For crystallization, we used a screening system [12] with ATP ϩ P i ϩ pyruvate < phosphoenolpyruvate ϩ AMP ϩ PP i hanging drops. Crystals grew only with one of the enzyme prepwhere phosphoenolpyruvate fixes CO 2 according to the arations and only at one condition, where the drop was a 1 :1 following reaction: mixture of 7 mg/ml enzyme with 2 mM Ap 5 A in 10 mM Hepes, pH 7.5 and 1 M Li 2 SO 4 with about 10% polyethylene glycolphosphoenolpyruvate ϩ CO 2 < oxaloacetate ϩ P i .8000. The latter solution was also used as the reservoir, which Among the about 50 known sequences of this kinase family was not completely air-tight, however, when the crystals apthere are only two from plants: rice [8] and maize [9]. The AKM peared. X-ray data were collected at room temperature using a sequence finds its closest homologues (42% identity) in Escheri-multi-wire ...

Mapping the Dynamics Landscape of Conformational Transitions in Enzyme: The Adenylate Kinase Case

“…References (73)(74)(75)(76)(77)(78)(79)(80)(81)(82)(83)(84)(85)(86)(87) appear in the Supporting Material.…”

The Dominance of Symmetry in the Evolution of Homo-oligomeric Proteins

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