ATP-grasp domain

Figure 16.25. Domain Structure of Pyruvate Carboxylase. The ATP-grasp domain activates HCO3 and transfers CO2 to the biotin-binding domain. From there, the CO2 is transferred to pyruvate generated in the central domain.

Figure 17.14. Structure of Succinyl CoA Synthetase. The enzyme is composed of two subunits. The a subunit contains a Rossmann fold that binds the ADP component of CoA, and the (3 subunit contains a nucleotideactivating region called the ATP-grasp domain. The ATP-grasp domain is shown here binding a molecule of ADP. The histidine residue picks up the phosphoryl group from near the CoA and swings over to transfer it to the nucleotide bound in the ATP-grasp domain.

This reaction takes place in a second ATP-grasp domain within the enzyme. The active sites leading to carbamic acid formation and carbamoyl phosphate formation are very similar, revealing that this enzyme evolved by a gene duplication event. Indeed, duplication of a gene encoding an ATP-grasp domain followed by specialization was central to the evolution of nucleotide biosynthetic processes (Section 25.2.3). 

Figure 25.3. Structure of Carbamoyl Phosphate Synthetase. This enzyme consists of two chains. The smaller chain (yellow) contains a site for glutamine hydrolysis to generate ammonia. The larger chain includes two ATP-grasp domains (blue and red). In one ATP-grasp domain (blue), bicarbonate is phosphorylated to carboxyphosphate, which then reacts with ammonia to generate carbamic acid. In the other ATP-grasp domain, the carbamic acid is phosphorylated to produce carbamoyl phosphate.

Nine additional steps are required to assemble the purine ring. Remarkably, the first six steps are analogous reactions. Most of these steps are catalyzed by enzymes with ATP-grasp domains that are homologous to those in carbamoyl phosphate synthetase. Each step consists of the activation of a carbon-bound oxygen atom (typically a... 

A few steps convert inosinate into either AMP or GMP (Figure 25,9). Adenylate is synthesized from inosinate by the substitution of an amino group for the carbonyl oxygen atom at C-6. Again, the addition of aspartate followed by the elimination of fumarate contributes the amino group. GXP, rather than ATP, is the phosphoryl-group donor in the synthesis of the adenylosuccinate intermediate from inosinate and aspartate. In accord with the use of GTP, the enzyme that promotes this conversion, adenylsuccinate synthase, is structurally related to the G-protein family and does not contain an ATP-grasp domain. The same enzyme catalyzes the removal of fumarate from adenylosuccinate in the synthesis of adenylate and from 5-aminoimidazole-4-jV-succinocarboxamide ribonucleotide in the synthesis of inosinate. 

Pyruvate carboxylase is of special interest because of its structural, catalytic, and allosteric properties. The N-terminal 300 to 350 amino acids form an ATP-grasp domain (Figure 16.23), which is a widely used ATP-activating... 

The active site for this reaction lies in a domain formed by the aminoterminal third of CPS. This domain forms a structure, called an ATP-grasp fold, that surrounds ATP and holds it in an orientation suitable for nucleophilic attack at the Y phosphoryl group. Proteins containing ATP-grasp folds catalyze the formation of carbon-nitrogen bonds through acyl-phosphate intermediates and are widely used in nucleotide biosynthesis. In the final step catalyzed by carbamoyl phosphate synthetase, carbamic acid is phosphorylated by another molecule of ATP to form carbamoyl phosphate. 

Each ACC half-reaction is catalyzed by a different protein sub-complex. The vitamin biotin is covalently coupled through an amide bond to a lysine residue on biotin carboxyl carrier protein (BCCP, a homodimer of 16.7-kDa monomers encoded by accB) by a specific enzyme, biotin-apoprotein ligase (encoded by birA), and is essential to activity. The crystal and solution structures of the biotinyl domain of BCCP have been determined, and reveal a unique thumb required for activity (J. Cronan, 2001). Carboxylation of biotin is catalyzed by biotin carboxylase (encoded by accC), a homodimeric enzyme composed of 55-kDa subunits that is copurified complexed with BCCP. The accB and accC genes form an operon. The three-dimensional structure of the biotin carboxylase subunit has been solved by X-ray diffraction revealing an ATP-grasp motif for nucleotide binding. The mechanism of biotin carboxylation involves the reaction of ATP and CO2 to form the shortlived carboxyphosphate, which then interacts with biotin on BCCP for CO2 transfer to the I -nitrogen. 

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