Structure-function relationships in a type I antifreeze polypeptide. The role of threonine methyl and hydroxyl groups in antifreeze activity.
Several analogs of an alanine-rich, alpha-helical type I antifreeze polypeptide from the winter flounder were synthesized and studied to evaluate the role of threonine residues on antifreeze activity. In this series, the four Thr residues in the wild type polypeptide were substituted with from one to four Ser, allo-Thr, or Val residues. Circular dichroism studies determined that these substitutions did not significantly diminish alpha-helicity. Thermal hysteresis data showed that substitution of Thr by Ser resulted in moderate to complete loss of antifreeze activity, depending on the number and position of the substituted Thr residue(s). Replacement by Val, in confirmation of other recent reports, or by allo-Thr had a much less detrimental effect on activity though there were qualitative differences in activity between the mutants and the wild type AFP. Based on these results, we propose that both the methyl and hydroxyl groups of Thr, particularly of the central two Thr residues, Thr13 and Thr24, play key roles in the ice-binding properties of the antifreeze peptide. Specifically, the methyls participate in hydrophobic interactions with ice, which provide the driving force for binding and stability, whereas the hydroxyls and other polar residues control binding specificity and impart additional stability through hydrogen bonding.
Zhang
W
W
Department of Chemistry, Boston University, Boston, Massachusetts 02215, USA.
Laursen
R A
RA
D000595:Amino Acid Sequence; D000818:Animals; D021301:Antifreeze Proteins; D001665:Binding Sites; D005399:Fishes; D005615:Freezing; D006023:Glycoproteins / Q000737:chemistry*; D007053:Ice; D008969:Molecular Sequence Data; D011489:Protein Denaturation; D017433:Protein Structure, Secondary; D013329:Structure-Activity Relationship; D013912:Threonine / Q000737:chemistry*
D021301:Antifreeze Proteins; D006023:Glycoproteins; D007053:Ice; D013912:Threonine