Crystal structure and substrate binding modeling of the uroporphyrinogen-III decarboxylase from Nicotiana tabacum - Implications for the catalytic mechanism

Humboldt-Universität zu Berlin, Berlín, Berlin, Germany
Journal of Biological Chemistry (Impact Factor: 4.57). 12/2001; 276(47):44108-16. DOI: 10.1074/jbc.M104759200
Source: PubMed


The enzymatic catalysis of many biological processes of life is supported by the presence of cofactors and prosthetic groups
originating from the common tetrapyrrole precursor uroporphyrinogen-III. Uroporphyrinogen-III decarboxylase catalyzes its
conversion into coproporphyrinogen-III, leading in plants to chlorophyll and heme biosynthesis. Here we report the first crystal
structure of a plant (Nicotiana tabacum) uroporphyrinogen-III decarboxylase, together with the molecular modeling of substrate binding in tobacco and human enzymes.
Its structural comparison with the homologous human protein reveals a similar catalytic cleft with six invariant polar residues,
Arg32, Arg36, Asp82, Ser214 (Thr in Escherichia coli), Tyr159, and His329 (tobacco numbering). The functional relationships obtained from the structural and modeling analyses of both enzymes allowed
the proposal for a refined catalytic mechanism. Asp82 and Tyr159 seem to be the catalytic functional groups, whereas the other residues may serve in substrate recognition and binding, with
Arg32 steering its insertion. The crystallographic dimer appears to represent the protein dimer under physiological conditions.
The dimeric arrangement offers a plausible mechanism at least for the first two (out of four) decarboxylation steps.

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Available from: Albrecht Messerschmidt, Jan 06, 2016
    • "Initially, chemical and biochemical analysis shaped the first period of the research on tetrapyrroles and their synthesis , before molecular and genetic analysis helped to identify genes and elucidated transcriptional control of TBS. Presently, the research moves towards biochemistry and structural analyses of the 3D structures of TBS proteins [16] [17] [18] [19], which subsequently lead to the exploration of multi-enzymatic protein complexes in vivo [20] [21] [22] [23] [24], as well as the elucidation of a potent network of metabolic and regulatory interactions [25] [26]. Some of these complexes and networks have been proposed years ago [27] [28] [29], but the characterization of the physical interactions between particular components still requires elucidation. "
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    ABSTRACT: Tetrapyrroles are macrocyclic molecules with various structural variants and multiple functions in Prokaryotes and Eukaryotes. Present knowledge about the metabolism of tetrapyrroles reflects the complex evolution of the pathway in different kingdoms of organisms, the complexity of structural and enzymatic variations of enzymatic steps, as well as a wide range of regulatory mechanisms, which ensure adequate synthesis of tetrapyrrole end-products at any time of development and environmental condition. This review intends to highlight new findings of research on tetrapyrrole biosynthesis in plants and algae. In the course of the heme and chlorophyll synthesis in these photosynthetic organisms, glutamate, one of the central and abundant metabolites, is converted into highly photoreactive tetrapyrrole intermediates. Thereby, several mechanisms of posttranslational control are thought to be essential for a tight regulation of each enzymatic step. Finally, we wish to discuss the potential role of tetrapyrroles in retrograde signaling and point out perspectives of the formation of macromolecular protein complexes in tetrapyrrole biosynthesis as an efficient mechanism to ensure a fine-tuned metabolic flow in the pathway. This article is part of a Special Issue entitled: Chloroplast Biogenesis. Copyright © 2015. Published by Elsevier B.V.
    No preview · Article · May 2015 · Biochimica et Biophysica Acta
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    • "The proteins of photosynthesis and carbon assimilation are represented by the alpha subunit of ATP synthase (spots U7 and U15), that catalyzes ATP synthesis/hydrolysis coupled with a transmembrane H + -translocation in chloroplasts and mitochondria [25]; rubisco activase (spot D12), which promotes and maintains the catalytic activity of Rubisco [26]; and uroporphyrinogen decarboxylase (spot D13), which is located at the first branch point of the ubiquitous tetrapyrrole biosynthetic pathway. Decarboxylation of uroporphyrinogen leads to the biosynthesis of hemes and chlorophylls, whereas its C-methylation initiates the synthesis of vitamin B 12 , sirohemes, and the nickel-chelating cofactor F 430 [27]. Fig. 1. "
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    ABSTRACT: Comparison of protein expression in necrotic leaves and in normal leaves of wheat (Triticum aestivum L.) showed that the abundance of 39 proteins was changed significantly, and 26 of these proteins were identified. Analysis of the function of the differentially expressed proteins in the necrotic hybrid leaves showed that the cytoprotective heat shock proteins may be induced to maintain the integrity of other proteins, facilitating the intercellular transportation of vital cellular enzymes upon necrosis. The increased abundance of NADH dehydrogenase indicated that the chloroplasts of necrotic leaves were under photo-oxidative stress. In addition, the light and dark events of photosynthesis were impacted differently during necrosis. The increased abundance of the hormone-sensitive enzymes phospholipase and β-1,3-glucanase suggested that the level of plant hormones may be increased in necrotic leaves. Both DNA helicase and maturase K were down-regulated in necrotic leaves, indicating basic genetic processes, including replication, repair, recombination, transcription and translation, were impacted during necrosis. The results of this study give a comprehensive picture of the post-transcriptional response to necrosis in hybrid wheat leaves and serve as a platform for further characterization of gene function and regulation in wheat hybrid necrosis.
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    • "However, a dimeric structure was postulated in the case of chicken (Seki et al. 1986), humans (Whitby et al. 1998), and Nicotiana tabacum (Martins et al. 2001), suggesting that dimer formation would be a common property of the UroD protein family. In addition, a number of biochemical and structural studies propose the existence of a dimer-dependent catalysis (Martins et al. 2001). Species from the genus Chlorella are among the most widely distributed microalgae, and thus are found in phytoplankton from all kinds of environments. "
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    ABSTRACT: Uroporphyrinogen decarboxylase (UroD) (EC is an enzyme from the tetrapyrrole biosynthetic pathway, in which chlorophyll is the main final product in algae. This is the first time that a study on UroD activity has been performed in a green alga (Chlorella). We isolated and partially purified the enzyme from a Chlorella kessleri (Trebouxiophyceae, Chlorophyta) strain (Copahue, Neuquén, Argentina), and describe for the first time some of its properties. In C. kessleri, the decarboxylation of uroporphyrinogen III occurs in two stages, via 7 COOH and then 6 and 5 COOH intermediates, with the decarboxylation of the 7 COOH compound being the rate-limiting step for the reaction. Cultures in the exponential growth phase showed the highest specific activity values. The most suitable conditions to measure UroD activity in C. kessleri were as follows: 0.23-0.3 mg protein/mL, approximately 6-8 micromol/L uroporphyrinogen III, and 20 min incubation time. Gel filtration chromatography and Western blot assays indicated that UroD from C. kessleri is a dimer of approximately 90 kDa formed by species of lower molecular mass, which conserves enzymatic activity.
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