Sirtuin is a protein with an enzymatic activity of NAD + -dependent protein deacetylation. It was first identified in yeast and its homologous genes have been widely found in various organisms. In bacteria, sirtuin gene was first described as cobB, encoding a cobalamin processing enzyme; and later its potential involvement in regulating acetylation levels of metabolic enzymes, transcription factors, chemotactic proteins and others have been reported. In order to study its physiological relevance in probiotic lactic acid bacteria, we analyzed the whole genome of three L. paracasei strains. All strains tested had sirtuin homolog genes designated hereby as sirA, and one of them had an additional gene designated as sirB. Following confirmation of their coding sequences by individual gene cloning, corresponding recombinant proteins have been generated and purified. The enzymatic characterization revealed that the intrinsic NAD + -dependent deacetylation activity of LpSirA (protein encoded by sirA) is comparable to human SIRT1. Furthermore, by blocking sirtuin activity using nicotinamide in vivo, together with an in vitro deacetylation reaction using recombinant LpSirA, we identified one of the target proteins in the lactic acid bacteria as the 30S ribosomal protein S4 (rpsD product).
We report here the draft genome sequences of three strains of Lactobacillus paracasei (NRIC 0644, NRIC 1781, and NRIC 1917) isolated from different sources. The three genomes range from 2.95 to 3.15 Mb with a G+C content of 46% and contain approximately 2,700 protein coding sequences.
Sirtuin has been associated in prolonging lifespan of different model organisms. It has been shown to have an enzymatic activity of NAD-dependent protein deacetylation targeting acetylated proteins. To determine targets and possible roles of sirtuin (LpSirA) in the Lactobacillus paracasei BL23, deletion (ΔsirA), sirtuin overexpressor (highsirA) and GFP fusion (highsirA-Venus) strains were generated, and microscopic localization and cell length analysis were done. Microscopic analysis revealed localization of LpSirA at cell division plates, at cell poles and all throughout the cell length in a spiral manner. Cell length analysis revealed that 46.9% of the ΔsirA cells were observed to be shorter (<2 μm), whereas 12.6% of the highsirA cells were observed to be longer (>4 μm) in comparison with the wild-type with only 17.1% short cells and 5.3% long cells. Our results suggest that sirtuin may have an essential role in cell division and cell shape regulation.
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