STRUCTURE-FUNCTION-RELATIONSHIPS IN AN ANTIFREEZE POLYPEPTIDE - THE EFFECT OF ADDED BULKY GROUPS ON ACTIVITY

We have proposed that the antifreeze activity of an alanine-rich alpha -helical antifreeze polypeptide (AFP) is due in part to side-by-side h ydrophobic interactions of AFP molecules bound on the ice surface (Wen , D., and Laursen, R. A. (1992b) Biophys. J. 63, 1659-1662). To test t his hypothesis and to assess the importance of a hydrophobic surface o n the outward facing (non-ice-binding) portions of the helix, we synth esized several AFP analogs with up to four Ala --> Gln or Ala --> Leu replacements and determined the effect of these changes on antifreeze activity and helix stability. Although Gln replacements caused some he lix destabilization and resultant reduction of antifreeze activity, th ey were in general well tolerated, suggesting that the bulk hydrophobi city of the non-ice-binding faces of the AFP, per se, is probably not a major factor determining AFP activity. However, placement of either Gln or Leu in position 17 on one side of the helix completely abolishe d activity, demonstrating that the specific location of bulky groups c an dramatically alter activity. We conclude from molecular modeling st udies that the effect seen by placing bulky groups in position 17 is d ue to steric hindrance that prevents effective association or packing of AFP molecules on the ice surface, in support of our hypothesis.