Molecular dynamics simulations elucidate the misfolding mechanisms of secretion-defective pancreatic lipase variants

Protein misfolding of pancreatic secretory enzymes is increasingly recognized as a contributor to chronic pancreatitis. Recently, missense pancreatic lipase (PNLIP) mutations that impair lipase secretion and cause intracellular aggregation and endoplasmic reticulum (ER) stress have been linked to the disease. Among these, A174P, G233E, and C254R are located in the N-terminal domain, whereas V454F is situated in the C-terminal domain of PNLIP. To elucidate how these mutations disrupt PNLIP structure, we performed detailed molecular dynamics simulations and structural modeling of human PNLIP variants. Evolutionary conservation analyses suggested that the mutation sites contribute to PNLIP structural stability. All mutations, regardless of location, altered the residue interaction network within the N-terminal PNLIP domain, consistently affecting Glu270. Detailed structural analysis revealed that the A174P substitution disrupted several helix-stabilizing hydrogen bonds, whereas the G233E mutation introduced multiple local hydrogen bonds and caused structural distortions. The C254R mutation abolished the native Cys254-Cys278 disulfide bond and instead formed an alternative salt bridge destabilizing the lid region. All three mutations altered stabilizing interactions mainly within the N-terminal domain of PNLIP. Interestingly, the V454F substitution exerted long-range allosteric effects, modulating key interactions also within the N-terminal domain. Further structural analysis showed that Phe454 in the variant became solvent-exposed, potentially leading to aggregate formation through the mutation site. These observations provide a molecular basis for PNLIP misfolding and its downstream proteotoxic effects, supporting the development of targeted therapies for chronic pancreatitis.