Bacillus subtilis α-amylase: interactions of a partially folded conformer with small unilamellar vesicles

摘要

We studied the interactions between conformers of exocellular α-amylase and small unilamellar vesicles (SUV) composed of the major membrane lipids of Bacillus subtilis under physiological conditions of pH, temperature and ionic strength. Using fluorescence spectroscopy, surface plasmon resonance (SPR) and phase separation, we show that the native α-amylase has no affinity for the SUV, whereas a partially folded form, displaying structural properties in common with the competent state for secretion, binds to the vesicles (K_A≈105 M~(-1)). This association prevented its subsequent folding. The complex was destabilized in the presence of PrsA, a major peripheric lipoprotein of B. subtilis which displays a strong affinity for SUV (KA≈1.5 * 10~8 M~(-1)). Vesicles coated with PrsA lost their ability to bind the partially folded conformer. The approach in vitro, in which our aim was to mimic the last stage of α-amylase translocation, indicates that PrsA possibly helps, in vivo, the secreted protein to acquire its native conformation by modulating the interaction between the latter and the lipid polar heads on the trans side of the cytoplasmic membrane.