Tieking Markus, Ehrmann Matthias A, Vogel Rudi F, Gänzle Michael G
TU München, Lehrstuhl Technische Mikrobiologie, Weihenstephaner Steig 16, 85350, Freising, Germany.
Appl Microbiol Biotechnol. 2005 Mar;66(6):655-63. doi: 10.1007/s00253-004-1773-5. Epub 2004 Nov 17.
Exopolysaccharides (EPS) produced in situ by sourdough lactobacilli affect rheological properties of dough as well as bread quality and may serve as prebiotics. The aim of this study was to characterize EPS-formation by Lactobacillus sanfranciscensis TMW 1.392 at the molecular level. A levansucrase gene from L. sanfranciscensis TMW 1.392 encompassing 2,300 bp was sequenced. This levansucrase is predicted to be a cell-wall associated protein of 879 amino acids with a relative molecular weight (M(R)) of 90,000. The levansucrase gene was heterologously expressed in Escherichia coli and purified to homogeneity. The recombinant enzyme exhibited transferase and hydrolase activities and produced glucose, fructose, 1-kestose and levan from sucrose; truncation of the N-terminal domain did not affect catalytic activity. Kestose formation was enhanced relative to fructose and levan formation by low temperature or high sucrose levels. During growth in wheat doughs, strain TMW 1.392 utilized sucrose to form fructose, 1-kestose, and fructan, whereas a levansucrase deletion mutant, L. sanfranciscensis TMW 1392Deltalev, lost the ability to hydrolyze sucrose, and did not produce fructan or 1-kestose. These results indicate that, in L. sanfranciscensis TMW 1.392, sucrose metabolism and formation of fructan and 1-kestose is dependent on the activity of a single enzyme, levansucrase.
酸面团乳酸菌原位产生的胞外多糖（EPS）会影响面团的流变学特性以及面包品质，并且可能用作益生元。本研究的目的是在分子水平上表征旧金山乳杆菌TMW 1.392的EPS形成过程。对来自旧金山乳杆菌TMW 1.392的一个包含2300 bp的果聚糖蔗糖酶基因进行了测序。预测该果聚糖蔗糖酶是一种与细胞壁相关的蛋白质，含879个氨基酸，相对分子质量（M(R)）为90,000。该果聚糖蔗糖酶基因在大肠杆菌中进行了异源表达并纯化至均一。重组酶表现出转移酶和水解酶活性，能从蔗糖产生葡萄糖、果糖、蔗果三糖和果聚糖；N端结构域的截短不影响催化活性。低温或高蔗糖水平会使蔗果三糖的形成相对于果糖和果聚糖的形成得到增强。在小麦面团中生长期间，菌株TMW 1.392利用蔗糖形成果糖、蔗果三糖和果聚糖，而果聚糖蔗糖酶缺失突变体旧金山乳杆菌TMW 1392Deltalev失去了水解蔗糖的能力，并且不产生果聚糖或蔗果三糖。这些结果表明，在旧金山乳杆菌TMW 1.392中，蔗糖代谢以及果聚糖和蔗果三糖的形成依赖于单一酶即果聚糖蔗糖酶的活性。
