Department of Food, Bioprocessing and Nutrition Sciences, North Carolina State University, Raleigh, NC 27695, USA.
Microbiology (Reading). 2010 Nov;156(Pt 11):3360-3367. doi: 10.1099/mic.0.043158-0. Epub 2010 Sep 9.
Structural components of the cell surface have an impact on some of the beneficial attributes of probiotic bacteria. In silico analysis of the L. acidophilus NCFM genome sequence revealed the presence of a putative cell surface protein that was predicted to be a myosin cross-reactive antigen (MCRA). As MCRAs are conserved among many probiotic bacteria, we used the upp-based counterselective gene replacement system, designed recently for use in L. acidophilus, to determine the functional role of this gene (LBA649) in L. acidophilus NCFM. Phenotypic assays were undertaken with the parent strain (NCK1909) and deletion mutant (NCK2015) to assign a function for this gene. The growth of NCK2015 (ΔLBA649) was reduced in the presence of lactate, acetate, porcine bile and salt. Adhesion of NCK2015 to Caco-2 cells was substantially reduced for both stationary-phase (∼45 % reduction) and exponential-phase cells (∼50 % reduction). Analysis of NCK2015 by scanning electron microscopy revealed a longer cell morphology after growth in MRS broth compared to NCK1909. These results indicate a role for LBA649 in stress tolerance, cell wall division and adherence to Caco-2 cells.
细胞表面的结构成分对益生菌的一些有益特性有影响。对嗜酸乳杆菌 NCFM 基因组序列的计算机分析显示，存在一种假定的细胞表面蛋白，预测其为肌球蛋白交叉反应抗原（MCRA）。由于 MCRAs 在许多益生菌中都保守，我们使用了最近专为嗜酸乳杆菌设计的 upp 基的反向选择基因替换系统，以确定该基因（LBA649）在嗜酸乳杆菌 NCFM 中的功能作用。对亲本菌株（NCK1909）和缺失突变体（NCK2015）进行表型分析，以确定该基因的功能。在存在乳酸盐、醋酸盐、猪胆盐和盐的情况下，NCK2015（ΔLBA649）的生长受到抑制。NCK2015 对 Caco-2 细胞的粘附在静止期（减少约 45%）和指数生长期（减少约 50%）均显著降低。通过扫描电子显微镜对 NCK2015 进行分析，发现与 NCK1909 相比，在 MRS 肉汤中生长后细胞形态更长。这些结果表明 LBA649 参与了应激耐受、细胞壁分裂和与 Caco-2 细胞的粘附。
