Blackman Steve A, Smith Thomas J, Foster Simon J
Department of Molecular Biology and Biotechnology, University of Sheffield, Firth Court, Western Bank, Sheffield $10 2TN, UK.
Microbiology (Reading). 1998 Jan;144 ( Pt 1):73-82. doi: 10.1099/00221287-144-1-73.
A set of isogenic mutants of Bacillus subtilis 168, insertionally inactivated in the genes encoding a number of lytic enzymes and a sigma factor (sigma D, which controls the expression of a number of autolysins) was constructed. Phenotypic analysis of the mutants determined the individual and combined roles of the autolysins in vegetative growth. The major vegetative autolysins of B. subtilis, LytC (50 kDa amidase) and LytD (90 kDa glucosaminidase), were shown to have roles in cell separation, cell wall turnover, antibiotic-induced lysis and motility. LytC was also shown to have a role in general cell lysis induced by sodium azide. Renaturing SDS-PAGE of cell-wall-binding protein extracts of the mutant strains revealed the presence of a novel autolysin that was previously masked by LytC. This 49 kDa enzyme was shown to be sigma D-controlled and was identified as a candidate cell separation and cell wall turnover enzyme. A multiple mutant strain, lacking LytC, LytD and the 49 kDa enzyme, retained at least ten bands of autolytic activity. These may correspond to individual or proteolytically processed novel autolysins, the functions of which are unknown. The multiple mutant strains facilitate the study of these, and other lytic enzymes, to determine their cellular functions.
构建了一组枯草芽孢杆菌168的同基因突变体，这些突变体在编码多种裂解酶和一种σ因子（控制多种自溶素表达的σD）的基因中发生了插入失活。对这些突变体的表型分析确定了自溶素在营养生长中的个体作用和联合作用。枯草芽孢杆菌的主要营养自溶素LytC（50 kDa酰胺酶）和LytD（90 kDa氨基葡萄糖苷酶）在细胞分离、细胞壁周转、抗生素诱导的裂解和运动性中发挥作用。LytC还被证明在叠氮化钠诱导的一般细胞裂解中起作用。对突变菌株细胞壁结合蛋白提取物进行复性SDS-PAGE分析，发现了一种新的自溶素，该自溶素以前被LytC掩盖。这种49 kDa的酶被证明受σD调控，并被鉴定为细胞分离和细胞壁周转酶的候选酶。一个缺失LytC、LytD和49 kDa酶的多重突变菌株保留了至少十条自溶活性带。这些可能对应于单个或经蛋白水解加工的新型自溶素，其功能尚不清楚。多重突变菌株有助于研究这些裂解酶和其他裂解酶，以确定它们的细胞功能。
