Screening of cutaneous secretions from Leptodactylus vastus Lutz, 1930 (Anura: Leptodactylidae): identification of bioactive proteins and anticoagulant activity

Leptodactylus vastus, popularly known as the pepper frog or "jia," is an anuran species endemic to Brazil and found throughout all states of the Northeast region. This study evaluated the protein composition and biological activities of a L. vastus cutaneous gland secretion (LVS). The protein profile was characterized by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and mass spectrometry, followed by screening for lectins, proteases, trypsin-like enzymes, and trypsin inhibitors. LVS was also evaluated for hemolytic and coagulation-modulatory activities, as well as for antimicrobial activity against pathogenic bacteria and yeasts. SDS-PAGE revealed seven polypeptide bands, and mass spectrometry indicated that three of them share similarity with methanethiol oxidase (a selenium-binding protein) previously described in other frog species. Lectins and trypsin inhibitors were not detected in LVS. Zymography indicated the presence of various proteases; however, trypsin-like enzymes were absent. LVS exhibited low hemolytic activity (maximum 8.6%) and prolonged prothrombin time (PT) (control: 12.3 s) and activated partial thromboplastin time (aPTT) (control: 32.0 s), reaching 90 and 54 s, respectively, at the highest sample concentration (100 μg/mL). LVS did not display antibacterial or antifungal activity. In conclusion, the cutaneous secretion of L. vastus contains proteins similar to methanethiol oxidase and proteases, which may contribute to the anticoagulant activity affecting both the extrinsic and intrinsic pathways. This study serves as a starting point for the characterization and discovery of LVS-derived proteins with biomedical and biotechnological potential.