Domain V of m‐calpain shows the potential to form an oblique‐orientated α‐helix, which may modulate the enzyme's activity via interactions with anionic lipid

The activity of m‐calpain, a heterodimeric, Ca 2+ ‐dependent cysteine protease appears to be modulated by membrane interactions involving oblique‐orientated α‐helix formation by a segment, GTAMRILGGVI, in the protein's smaller subunit. Here, graphical and hydrophobic moment‐based analyses predicted that this segment may form an α‐helix with strong structural resemblance to the influenza virus peptide, HA2, a known oblique‐orientated α‐helix former. Fourier transform infrared spectroscopy showed that a peptide homologue of the GTAMRILGGVI segment, VP1, adopted low levels of α‐helical structure (≈ 20%) in the presence of zwitterionic lipid and induced a minor decrease (3 °C) in the gel to liquid‐crystalline phase transition temperature, T C , of the hydrocarbon chains of zwitterionic membranes, suggesting interaction with the lipid headgroup region. In contrast, VP1 adopted high levels of α‐helical structure (65%) in the presence of anionic lipid, induced a large increase (10 °C) in the T C of anionic membranes, and showed high levels of anionic lipid monolayer penetration (ΔSP = 5.5 mN·m −1 ), suggesting deep levels of membrane penetration. VP1 showed strong haemolytic ability (LD 50  = 1.45 m m ), but in the presence of ionic agents, this ability, and that of VP1 to penetrate anionic lipid monolayers, was greatly reduced. In combination, our results suggest that m‐calpain domain V may penetrate membranes via the adoption of an oblique‐orientated α‐helix and electrostatic interactions. We speculate that these interactions may involve snorkelling by an arginine residue located in the polar face of this α‐helix.