Structural basis of enzyme adaptation to alkaline pH

Abstract

pH is one of the key parameters that affect stability and function of proteins. The crystal structure of the vitamin B 6-dependent enzyme phosphoserine aminotransferase from the obligatory alkaliphile Bacillus alcalophilus has been determined at 1.08 A resolution [1], the highest resolution for any PLP-dependent enzyme so far. The final model contains 5824 protein atoms and 949 water molecules, and has an R-factor of 11.7% (Rfree = 13.9%) after anisotropic B-factor refinement with SHELX. The enzyme displays a narrow pH optimum of enzymatic activity at pH 9.0. The final structure was compared to the previously reported structure of themesophilic phosphoserine aminotransferase from Escherichia coli [2] and to that of phosphoserine aminotransferase froma facultative alkaliphile, Bacillus circulans subsp. alkalophilus [3]. All three enzymes are homodimers with each monomer comprising a two-domain architecture. Despite the high structural similarity, the alkaliphilic representatives possess a set of distinctive structural features. Two residues directly interacting with pyridoxal-5’-phosphate are replaced, and an additional hydrogen bond to theO3’ atomof the cofactor is present in alkaliphilic phosphoserine aminotransferases. The number of hydrogen bonds and hydrophobic interactions at the dimer interface is increased. Hydrophobic interactions between the two domains in the monomers are enhanced. Moreover, the number of negatively charged amino acid residues increases on the solvent accessible molecular surface and fewer hydrophobic residues are exposed to the solvent. Further, the total amount of ion pairs and ion networks is significantly reduced in theBacillus enzymes, while the total number of hydrogen bonds is increased. The mesophilic enzyme fromEscherichia coli contains two additional s-strands in a surface loop with a third s-strand being shorter in the structure. The identified structural features are proposed to be possible factors implicated in the alkaline adaptation of phosphoserine aminotransferase.