Exploring the role of a glycine cluster in cold adaptation of an alkaline phosphatase.
In an effort to explore the role of glycine clusters on the cold adaptation of enzymes, we designed point mutations aiming to alter the distribution of glycine residues close to the active site of the psychrophilic alkaline phosphatase from the Antarctic strain TAB5. The mutagenesis targets were residues Gly261 and Gly262. The replacement of Gly262 by Ala resulted in an inactive enzyme. Substitution of Gly261 by Ala resulted to an enzyme with lower stability and increased energy of activation. The double mutant G261A/Y269A designed on the basis of side-chain packing criteria from a modelled structure of the enzyme resulted in restoration of the energy of activation to the levels of the native enzyme and in an increased stability compared to the mutant G261A. It seems therefore, that the Gly cluster in combination with its structural environment plays a significant role in the cold adaptation of the enzyme.
Mavromatis
Konstantinos
K
Department of Biology, Division of Applied Biology and Biotechnology, University of Crete, Greece.
Tsigos
Iason
I
Tzanodaskalaki
Maria
M
Kokkinidis
Michael
M
Bouriotis
Vassilis
V
D000222:Adaptation, Physiological; D000469:Alkaline Phosphatase / Q000737:chemistry* / Q000378:metabolism; D000595:Amino Acid Sequence; D003080:Cold Temperature; D005998:Glycine; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D013329:Structure-Activity Relationship; D013816:Thermodynamics
D000469:Alkaline Phosphatase; D005998:Glycine