title: Studies of Glu-416 variants of beta-galactosidase (E. coli) show that the active site Mg(2+) is not important for structure and indicate that the main role of Mg (2+) is to mediate optimization of active site chemistry.
abstract: Variants of beta-galactosidase with Valine and with Glutamine replacing Glutamate-416 did not have a Mg(2+) bound at the active site even at high Mg(2+) concentrations (200 mM). They had low catalytic activity and the pH profiles were very different from those of the native enzyme. In addition, substrates, substrate analogs, transition state analogs and galactose bound very poorly. However, the orientation and conformation of the Mg(2+) ligands (residues 416, 418, and 461) as well as the B-factors of these three side chains did not change significantly. The structures, conformations and B-factors of other active site residues were also essentially unchanged. These studies show that the active site Mg(2+) is not necessary for structure and is, therefore, mainly important for modulating the chemistry and mediating the interactions between the active site components.
authors: Department of Biological Sciences, University of Calgary, Calgary, AB, T2N 1N4, Canada.
authors: Summie
authors: S
authors: Lo
authors: Megan L
authors: ML
authors: Dugdale
authors: Nisha
authors: N
authors: Jeerh
authors: Tabitha
authors: T
authors: Ku
authors: Nathan J
authors: NJ
authors: Roth
authors: Reuben E
authors: RE
authors: Huber
mesh_terms: D019943:Amino Acid Substitution; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D014644:Genetic Variation; D007700:Kinetics; D008274:Magnesium / Q000737:chemistry*; D008968:Molecular Conformation; D020125:Mutation, Missense; D001616:beta-Galactosidase / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism
chemical_list: D029968:Escherichia coli Proteins; D001616:beta-Galactosidase; D008274:Magnesium