Complete de novo sequencing of antimicrobial peptides in the venom of the scorpion Isometrus maculatus.

Scorpion venom contains antimicrobial peptides (AMPs) in addition to neurotoxic peptides. Recent extensive transcriptomic analysis of venom glands of various scorpion species dramatically increased the number of identified AMPs. However, identification of peptides from genetic information requires reference sequences of similar peptides, and moreover, it is often difficult to predict post-translational modifications. In this study, we searched for unknown AMPs contained in the Isometrus maculatus scorpion venom based on the structural features, such as the hydrophobic nature and the lack of disulfide bonds, which are commonly observed in the majority of scorpion AMPs. Their primary structures were determined only by a de novo sequencing method using two types of MS instruments, which induce peptide fragmentation in a different fashion. Chemical derivatization techniques were also used to facilitate Leu/Ile discrimination. As a result, total 15 AMP candidates were discovered from the I. maculatus venom. Among them, three peptides were identified as AMPs by evaluating their biological activity. Other 12 candidate peptides were structurally related to the identified AMPs, possibly generated by enzymatic cleavage of the mature peptides in the venom.