Trapping of a spiral-like intermediate of the bacterial cytokinetic protein FtsZ

The earliest stage in bacterial cell division is the formation of a ring, composed of the tubulin-like protein FtsZ, at the division site. Tight spatial and temporal regulation of Z-ring formation is required to ensure that division occurs precisely at midcell between two replicated chromosomes. However, the mechanism of Z-ring formation and its regulation in vivo remain unresolved. Here we identify the defect of an interesting temperature-sensitive ftsZ mutant (ts1) of Bacillus subtilis. At the nonpermissive temperature, the mutant protein, FtsZ(Ts1), assembles into spiral-like structures between chromosomes. When shifted back down to the permissive temperature, functional Z rings form and division resumes. Our observations support a model in which Z-ring formation at the division site arises from reorganization of a long cytoskeletal spiral form of FtsZ and suggest that the FtsZ(Ts1) protein is captured as a shorter spiral-forming intermediate that is unable to complete this reorganization step. The ts1 mutant is likely to be very valuable in revealing how FtsZ assembles into a ring and how this occurs precisely at the division site.

Evaluation of cross-linked enzyme aggregates of Lactobacillus cell-envelope proteinases, for protein degradation

Enzymatic hydrolysis is a widely used approach to improve the functional, nutritionaland physiological properties of food proteins. In this study, cross-linked enzyme aggre-gates (CLEAs) have been prepared from cell-envelope proteinases (CEPs) of Lactobacillusdelbrueckii subsp. lactis 313 and their proteolytic properties have been evaluated using severalfood proteins. We have optimized cross-linking conditions including ammonium sulphateconcentration, incubation temperatures, agitation speed, glutaraldehyde cross-linker con-centration, reaction time and the addition of proteic feeders. Particularly, the presence ofBSA improves retained activity of cross-linked CEP aggregates (CLCEPAs) from 21.5% to 40.9%.Blocking unreacted cross-linking groups on aggregates is important to enhance recyclabil-ity of CLCEPAs. CLCEPAs had attractive thermal stability at 50◦C and it showed enhancedcatalytic activity over long-term storage after lyophilization. We have demonstrated thatCLCEPAs has proteolytic properties on different food proteins including complex (chickenegg albumin, skimmed-milk protein), fractionated (bovine casein, whey protein isolate), andpurified (bovine serum albumin) proteins. Being the first report of CLEAs from lactobacilliCEPs, this study demonstrates the feasibility of using LDL 313 CLCEPAs for degradation ofvarious food proteins.