title: Isoleucine 259 and isoleucine 260 residues in Streptococcus gordonii soluble inorganic pyrophosphatase play an important role in enzyme activity.
abstract: The hinge region of the family II soluble inorganic pyrophosphatase (PPase) found in Streptococcus gordonii DL1 (Challis) has previously been shown to play an important role in opening and closing of the active site between the N- and C-terminal domains of PPase. Amino acid residues isoleucine 259 (I259) and isoleucine 260 (I260) are highly conserved among catalytically active family II PPases and are located very close to the hinge region. Substitution of either I259 or I260 with a hydrophilic acidic amino acid (glutamate or aspartate) resulted in adverse effects on the kinetic properties of the enzyme. The I259/E and I259/D variants were nearly catalytically inactive (k(cat)/K(m) <0.2% of the wild type), whereas both I260/E and I260/D variants showed less than 15% of the catalytic efficiency of the wild type S. gordonii PPase. Conservative substitution of both residues to valine (I259/V, I260/V) showed no significant effect on the catalytic activity. The solvent accessibility data for I259 and I260 and the proximity of these amino acids to the hinge region suggest that occlusion of these residues may stabilise the closed and open conformations of the protein, respectively, thus aiding the catalytic activity of the enzyme.
authors: School of Biosciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, UK. ilias14@yahoo.com
authors: Mohammad
authors: M
authors: Ilias
authors: Scott A
authors: SA
authors: White
authors: Tom W
authors: TW
authors: Young
mesh_terms: D001224:Aspartic Acid / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D002384:Catalysis; D020134:Catalytic Domain; D018698:Glutamic Acid / Q000235:genetics / Q000378:metabolism; D043564:Inorganic Pyrophosphatase / Q000235:genetics / Q000378:metabolism*; D007532:Isoleucine / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D054773:Streptococcus gordonii / Q000235:genetics / Q000378:metabolism*; D014633:Valine / Q000235:genetics / Q000378:metabolism
chemical_list: D007532:Isoleucine; D001224:Aspartic Acid; D018698:Glutamic Acid; D043564:Inorganic Pyrophosphatase; D014633:Valine