Lipid Transfer Proteins (LTP) with α-Amylase Inhibitory Activity from Capsicum chinense Seeds: Structural and Functional Insights

Lipid transfer proteins (LTPs) are small, cysteine-rich antimicrobial peptides widely distributed in plant tissues and involved in various physiological functions, especially defense against pathogens and pests. In Capsicum chinense, a species known for its richness in bioactive compounds, LTPs were previously identified in seeds, exhibiting antifungal and α-amylase inhibitory activities. Given this potential, the present study aimed to purify and structurally and functionally characterize the LTP-rich fraction (P10), previously isolated from C. chinense seeds. To this end, the P10 fraction was subjected to reversed-phase rechromatography, resulting in three subfractions (R1–R3) with molecular masses close to 9 kDa, which reacted positively with anti-LTP antibodies. The protein identities were confirmed by mass spectrometry, revealing nonspecific LTPs with high similarity to C. chinense reference sequences. AlphaFold structural modeling, combined with proteomic mapping, revealed conserved surface-exposed regions, suggesting important functional targets. Furthermore, circular dichroism analyses demonstrated the stability of the secondary structure of LTPs at different temperature ranges, as well as their ability to interact with model lipid membranes. Functional assays confirmed the significant inhibitory activity of these LTPs against Tenebrio molitor, human salivary, and swine pancreatic α-amylases, with minimal hemolytic effects. Corroborating these data, molecular docking studies revealed stable interactions between LTPs and target enzymes, with the participation of specific residues at the binding interface, suggesting a possible mechanism of enzyme inhibition. Taken together, the data obtained reinforce the multifunctional nature of LTPs from C. chinense seeds and highlight their promising biotechnological potential, especially for applications in agriculture and healthcare.