Genetic and proteomic analysis of factors affecting serum cholesterol reduction by Lactobacillus acidophilus A4
Appl Environ Microbiol. 2010 Jul;76(14):4829-35 doi: 10.1128/AEM.02892-09.
Abstract: This article identifies novel factors
involved in cholesterol reduction by probiotic bacteria, which were identified
using genetic and proteomic approaches. Approximately 600 Lactobacillus
acidophilus A4 mutants were created by random mutagenesis. The
cholesterol-reducing ability of each mutant was determined and verified using
two different methods: the o-phthalaldehyde assay and gas chromatographic
analysis (GC). Among screened mutants, strain BA9 showed a dramatically
diminished ability to reduce cholesterol, as demonstrated by a 7.7% reduction
rate, while the parent strain had a more than 50% reduction rate. The transposon
insertion site was mapped using inverse PCR (I-PCR), and it was determined using
bioinformatic methods that the deleted region contained the Streptococcus
thermophilus catabolite control protein A gene (ccpA). In addition, we have
shown using two-dimensional gel electrophoresis (2-DE) that several proteins,
including a transcription regulator, FMN-binding protein, major facilitator
superfamily permease, glycogen phosphorylase, the YknV protein, and
fructose/tagatose bisphosphate aldolase, were strongly regulated by the ccpA
gene. In addition, in vivo experiments investigating ccpA function were
conducted with rats. Rats fed wild-type L. acidophilus A4 showed a greater than
20% reduction in total serum cholesterol, but rats fed BA9 mutant L. acidophilus
showed only an approximately 10% reduction in cholesterol. These results provide
important insights into the mechanism by which these lactic acid bacteria reduce
cholesterol.