THE CONTRIBUTION OF THE CELL-WALL TO A TRANSMEMBRANE CALCIUM GRADIENTCOULD PLAY A KEY ROLE IN BACILLUS-SUBTILIS PROTEIN SECRETION

A weak Ca2+-binding site (K(a) = 0.8 x 10(3) M-1, at pH 7) was identif ied in the mature part of levansucrase. An amino acid substitution (Th r-236-->Ile) in this site alters simultaneously the affinity for calci um, the folding transition and the efficiency of the secretion process of levansucrase. Moreover, the ability of the Bacillus subtilis cell wall to concentrate calcium ions present in the culture medium was stu died. We confirm the results of Beveridge and Murray who showed that t he concentration factor is about 100 to 120 times. This property prese rves a high concentration of Ca2+ (>2 mM) on the external side of the cytoplasmic membrane, even in the absence of further Ca2+ supplementat ion in the growth medium. Such local conditions allow the spontaneous unfolding-folding transition of levansucrase en route for secretion. S ince several exocellular proteins of B. subtilis are calcium-binding p roteins, we propose that the high concentration of calcium ion in the microenvironment of the cell wall may play a key role in the ultimate step of their secretion process.