Heman Chao

New ice‐binding face for type I antifreeze protein

Type I antifreeze protein (AFP) from winter flounder is an alanine‐rich, 37 amino acid, single α‐helix that contains three 11 amino acid repeats (Thr‐X2‐Asx‐X7), where X is generally Ala. The regularly spaced Thr, Asx and Leu residues lie on one face of the helix and have traditionally been thought to form hydrogen bonds and van der Waals interactions with the ice surface. Recently, substitution experiments have called into question the importance of Leu and Asn for ice‐binding. Sequence alignments of five type I AFP isoforms show that Leu and Asn are not well conserved, whereas Ala residues adjacent to the Thr, at right angles to the Leu/Asn‐rich face, are completely conserved. To investigate the role of these Ala residues, a series of Ala to Leu steric mutations was made at various points around the helix. All the substituted peptides were fully α‐helical and remained as monomers in solution. Wild‐type activity was retained in A19L and A20L. A17L, where the substitution lies adjacent to the Thr‐rich face, had no detectable antifreeze activity. The nearby A21L substitution had 10% wild‐type activity and demonstrated weak interactions with the ice surface. We propose a new ice‐binding face for type I AFP that encompasses the conserved Ala‐rich surface and adjacent Thr.

A new paradigm for fish antifreeze protein binding to ice

Michael Houston, Jr., Heman Chao, Michele C. Loewen, Frank D. Sonnichsen, Cyril M. Kay, Brian D. Sykes, Peter L. Davies and Robert S. Hodges Protein Engineering Network of Centres of Excellence, University of Alberta, Edmonton, Alberta, T6G 2S2, Canada. Department of Biochemistry, Queen’s University, Kingston, Ontario, K7L 3N6, Canada; Department of Physiology and Biophysics, Case Western Reserve University, Cleveland, OH 44106-4970, USA