Overproduction of a dextranase inhibitor by Streptococcus sobrinus mutants.
Overproduction of a dextranase inhibitor by Streptococcus sobrinus mutants.
复制标题
远缘链球菌突变体过量产生葡聚糖酶抑制剂。
DOI：
10.1128/jb.176.23.7206-7212.1994
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发表时间：
1994
影响因子：
3.2
通讯作者：
Curtiss3rd,R
中科院分区：
文献类型：
--
作者：
Wanda,SY;Camilli,A;Murchison,HM;Curtiss3rd,R
在甲基磺酸乙酯诱变后鉴定的UAB66突变体的细胞外部分中检测到一种sobrinus内酯葡聚糖酶抑制剂，该突变体要么缺乏葡聚糖酶活性（Dex-），要么过量产生水溶性葡聚糖。两组突变体具有相同的表型，并且在细胞外部分的检测中显示没有葡聚糖酶活性（H. Murchison， S. Larrimore和R. Curtiss III, infection）。（H. Murchison， S. Larrimore, and R. Curtiss III，《感染》，1981），并在体外共培养过程中显示出粘附缺陷（Adh-）和抑制野生型菌株粘附的能力。[j] .高田，薛田，李晓明，李晓明，《生物医学工程学报》。免疫学。50:833-843,1985)。通过免疫印迹（Western blots）和随后的蓝色葡聚糖-十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（BD-SDS-PAGE）分析，证明这些Dex-突变体确实合成了酶活性葡聚糖酶。混合实验的结果表明，这些Dex- Adh突变体产生了大量的细胞表面定位或细胞相关的葡聚糖酶抑制剂（Dei）。Dei对热稳定，但对胰蛋白酶敏感。在BD-SDS-PAGE后加入多余的葡聚糖酶，检测到Dei为蓝带，表观分子质量分别为43、40、37、27和23 kDa。Dei竞争性地抑制葡聚糖酶活性，由野生型S. sobrinus菌株合成，其数量取决于生长介质和生长周期的阶段。R. M. Hamelik和M. M. McCabe（生物化学）Biophys。Res. common . 106:875- 880,1982)先前描述了野生型sobrinus菌株中的Dei。
An inhibitor of Streptococcus sobrinus endodextranase was detected in the extracellular fractions of UAB66 mutants identified following ethyl methanesulfonate mutagenesis as either devoid of dextranase activity (Dex-) or overproducing water-soluble glucan. The two groups of mutants had the same phenotype and displayed no dextranase activity in assays of extracellular fractions (H. Murchison, S. Larrimore, and R. Curtiss III, Infect. Immun. 34:1044-1055, 1981) and had been shown to be defective in adherence (Adh-) and capable of inhibiting adherence of wild-type strains during cocultivation in vitro (H. Murchison, S. Larrimore, and R. Curtiss III, Infect. Immun. 50:826-832, 1985) and in vivo in gnotobiotic rats (K. Takada, T. Shiota, R. Curtiss III, and S. M. Michalek, Infect. Immun. 50:833-843, 1985). By analysis of proteins in Western blots (immunoblots) and following blue dextran-sodium dodecyl sulfate-polyacrylamide gel electrophoresis (BD-SDS-PAGE), it was demonstrated that these Dex- mutants did synthesize enzymatically active dextranase. From the results of mixing experiments, it was determined that these Dex- Adh- mutants produced enhanced amounts of a cell surface-localized or a cell-associated dextranase inhibitor (Dei). Dei was heat stable but trypsin sensitive. By adding excess dextranase following BD-SDS-PAGE, Dei was detected as blue bands with apparent molecular masses of 43, 40, 37, 27, and 23 kDa. Dei competitively inhibits dextranase activity and is synthesized by wild-type S. sobrinus strains, with the amount varying depending upon growth medium and stage in the growth cycle. R. M. Hamelik and M. M. McCabe (Biochem. Biophys. Res. Commun. 106:875-880, 1982) previously described a Dei in a wild-type S. sobrinus strain.