TableEdit

Yakushiji, Y, Nishikori, S, Yamanaka, K and Ogura, T (2006) Mutational analysis of the functional motifs in the ATPase domain of Caenorhabditis elegans fidgetin homologue FIGL-1: firm evidence for an intersubunit catalysis mechanism of ATP hydrolysis by AAA ATPases. J. Struct. Biol. 156:93-100

The AAA family proteins usually form a hexameric ring structure. The ATP-binding pocket, which is located at the interface of subunits in the hexamer, consists of three functionally important motifs, the Walker A and B motifs, and the second region of homology (SRH). It is well known that Walker A and B motifs mediate ATP binding and hydrolysis, respectively. Highly conserved arginine residues in the SRH have been proposed to function as arginine fingers, which interact with the gamma-phosphate of bound ATP. To elucidate the mechanism of ATP hydrolysis, we prepared several mutants of the Caenorhabditis elegans fidgetin homologue FIGL-1 carrying a mutation in each of the above-mentioned three motifs. None of the constructed mutants showed ATPase activity. All the mutants except for K362A were able to bind ATP. A decrease in the ATPase activity by mixing wild-type and each mutant subunits was caused by the formation of hetero-hexamers. Mixtures of E416A and R471A, or N461A and R471A led to the formation of hetero-hexamers with partially restored ATPase activities, providing direct, firm evidence for the intersubunit catalysis model. In addition, based on the results obtained with mixtures of K362A with wild-type or R471A subunits, we propose that a conformational change upon ATP binding is required for proper orientation of the arginine fingers, which is essential for efficient hydrolysis of ATP bound to the neighboring subunit.

PubMed Online version:10.1016/j.jsb.2006.03.001

Adenosine Triphosphatases/analysis; Adenosine Triphosphatases/chemistry; Adenosine Triphosphatases/genetics; Adenosine Triphosphatases/metabolism; Amino Acid Motifs; Amino Acid Substitution; Animals; Arginine/chemistry; Arginine/genetics; Arginine/metabolism; Caenorhabditis elegans Proteins/chemistry; Caenorhabditis elegans Proteins/genetics; Caenorhabditis elegans Proteins/isolation & purification; Caenorhabditis elegans Proteins/metabolism; Catalysis; Hydrolysis; Models, Biological; Models, Molecular; Nuclear Proteins; Protein Structure, Tertiary; Protein Subunits/chemistry; Protein Subunits/genetics; Protein Subunits/metabolism; Spodoptera/cytology