Synthesis and Order of Assembly of Spore Coat Proteins in Bacillus subtilis

Abstract

Analysis of mature spores or their integuments by extraction with sodium dodecyl sulphate/dithioerythritol followed by electrophoresis shows that the coat contains four major proteins and about ten others. Nine of the 14 proteins begin to be synthesized in stage II or stage III and their synthesis must be controlled by stage II or stage III operons. Some of these proteins are incorporated into the spore structure from about t 4 onwards (i.e. 4 h after induction of sporulation). Their deposition in the coat between t 4 and t 7 is not stopped by chloramphenicol, with the exception of one protein (mol. wt 36000) which begins to be synthesized only at t 6. Spores were isolated at various stages from about t 5 onwards, and the surface proteins were labelled with 125I. The labelling patterns show that proteins which are exposed on the surface at t 5·3 are successively overlaid as the spores mature. It appears that the coat of the mature spore contains at least three layers. The outermost layer is mainly composed of an alkali-soluble protein (mol. wt 12 000), which is synthesized early (t 2), and the 36000 mol. wt protein, which is synthesized very late. Deposition of the former seems to require processing by proteolytic action, and both proteins are apparently necessary for the acquisition of resistance to lysozyme, though not for resistance to heat or organic solvents. The results are discussed in relation to the classification of sporulation events and the nomenclature of the genetic loci controlling sporulation.