Dataset Information

Structure of Escherichia coli aspartate ?-decarboxylase Asn72Ala: probing the role of Asn72 in pyruvoyl cofactor formation.

ABSTRACT: The crystal structure of the Asn72Ala site-directed mutant of Escherichia coli aspartate ?-decarboxylase (ADC) has been determined at 1.7 Å resolution. The refined structure is consistent with the presence of a hydrolysis product serine in the active site in place of the pyruvoyl group required for catalysis, which suggests that the role of Asn72 is to protect the ester formed during ADC activation from hydrolysis. In previously determined structures of activated ADC, including the wild type and other site-directed mutants, the C-terminal region of the protein is disordered, but in the Asn72Ala mutant these residues are ordered owing to an interaction with the active site of the neighbouring symmetry-related multimer.

SUBMITTER: Webb ME

PROVIDER: S-EPMC3325809 | biostudies-literature | 2012 Apr

REPOSITORIES: biostudies-literature

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Structure of Escherichia coli aspartate α-decarboxylase Asn72Ala: probing the role of Asn72 in pyruvoyl cofactor formation.

Webb Michael E ME Lobley Carina M C CM Soliman Fatima F Kilkenny Mairi L ML Smith Alison G AG Blundell Tom L TL Abell Chris C

Acta crystallographica. Section F, Structural biology and crystallization communications 20120328 Pt 4

The crystal structure of the Asn72Ala site-directed mutant of Escherichia coli aspartate α-decarboxylase (ADC) has been determined at 1.7 Å resolution. The refined structure is consistent with the presence of a hydrolysis product serine in the active site in place of the pyruvoyl group required for catalysis, which suggests that the role of Asn72 is to protect the ester formed during ADC activation from hydrolysis. In previously determined structures of activated ADC, including the wild type and ...[more]

PMID: 22505409

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Project description:Enzymes catalyze a particular reaction in cells, but only a few control the rate of this reaction and the metabolic pathway that follows. One specific mechanism for such enzymatic control of a metabolic pathway involves molecular feedback, whereby a metabolite further down the pathway acts at a unique site on the control enzyme to alter its activity allosterically. This regulation may be positive or negative (or both), depending upon the particular system. Another method of enzymatic control involves the cooperative binding of the substrate, which allows a large change in enzyme activity to emanate from only a small change in substrate concentration. Allosteric regulation and homotropic cooperativity are often known to involve significant conformational changes in the structure of the protein. Escherichia coli aspartate transcarbamoylase (ATCase) is the textbook example of an enzyme that regulates a metabolic pathway, namely, pyrimidine nucleotide biosynthesis, by feedback control and by the cooperative binding of the substrate, L-aspartate. The catalytic and regulatory mechanisms of this enzyme have been extensively studied. A series of X-ray crystal structures of the enzyme in the presence and absence of substrates, products, and analogues have provided details, at the molecular level, of the conformational changes that the enzyme undergoes as it shifts between its low-activity, low-affinity form (T state) to its high-activity, high-affinity form (R state). These structural data provide insights into not only how this enzyme catalyzes the reaction between l-aspartate and carbamoyl phosphate to form N-carbamoyl-L-aspartate and inorganic phosphate, but also how the allosteric effectors modulate this activity. In this Account, we summarize studies on the structure of the enzyme and describe how these structural data provide insights into the catalytic and regulatory mechanisms of the enzyme. The ATCase-catalyzed reaction is regulated by nucleotide binding some 60 Å from the active site, inducing structural alterations that modulate catalytic activity. The delineation of the structure and function in this particular model system will help in understanding the molecular basis of cooperativity and allosteric regulation in other systems as well.

Dataset Information

Structure of Escherichia coli aspartate ?-decarboxylase Asn72Ala: probing the role of Asn72 in pyruvoyl cofactor formation.

Publications

Structure of Escherichia coli aspartate α-decarboxylase Asn72Ala: probing the role of Asn72 in pyruvoyl cofactor formation.

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