High Resolution Structures and Structure-Function Relationships in Histidine-Rich Antimicrobial Peptides from Cod

Abstract

Two paralogous antimicrobial peptide (AMP) sequences, Gad-1 and Gad-2, were previously identified from an Atlantic cod (Gadus morhua) expressed sequence tag database. Both peptides are rich in histidine, suggesting that their activity might be pH dependent. Indeed, minimal inhibitory concentration (MIC) assays with Gram-negative bacteria demonstrate that the activity of Gad-2 is substantially higher at pH 5 than it is at pH 7, whereas the activity of Gad-1 at pH 5 is similar to its activity at pH 7. The paralogues also appear to differ from each other in their level of activity against Gram-negative bacteria, with Gad-1 exhibiting more activity than Gad-2, even at pH 5. Clues to the origin of the different pH dependencies of the activity of the two peptides, as well as the difference in activity of Gad-1 and Gad-2 at pH 5, were provided by circular dichroism (CD) studies and high resolution NMR structures in sodium dodecyl sulfate (SDS) micelles at pH 5. Gad-1 takes on a helical configuration from residue 4 to 21. Gad-2 also appears to be predominantly helical but has a kink at the junction between H10 and H11. This deformation in the helix is likely due to the electrostatic repulsion between histidine sidechains at pH 5.