Salt stress represses production of extracellular proteases in Bacillus pumilus

Abstract

ABSTRACT.Bacillus pumilus is able to secrete subtilisin-like proteases, one of which has been purified and characterized biochemically, demonstrating great potential for use in industrial applications. In the current study, the biosynthesis and transcription of extracellular proteases in B. pumilus (BA06) under salt stress were investigated using various methods, including a proteolytic assay, zymogram analysis, and real-time PCR. Our results showed that total extracellular proteolytic activity, both in fermentation broth and on milk-containing agar plates, was considerably repressed by salt in a dosage-dependent manner. As Bacillus species usually secret multiple extracellular proteases, a variety of individual extracellular protease encoding genes were selected for real-time PCR analysis. It was shown that proteases encoded by the aprE and aprX genes were the major proteases in the fermentation broth in terms of their transcripts in B. pumilus. Further, transcription of aprE, aprX, and epr genes was indeed repressed by salt stress. In contrast, transcription of other genes (e.g., vpr and wprA) was not repressed or significantly affected by the salt. Conclusively, salt stress represses total extracellular proteolytic activity in B. pumilus, which can largely be ascribed to suppression of the major protease-encoding genes (aprE, R.F. Liu et al. 4940©FUNPEC-RP www.funpecrp.com.br Genetics and Molecular Research 14 (2): 4939-4948 (2015) aprX) at the transcriptional level. In contrast, transcription of other protease-encoding genes (e.g., vpr, wprA) was not repressed by salt stress.

Transcriptome profiling analysis reveals metabolic changes across various growth phases in Bacillus pumilus BA06

“…Fig. 1b shows that the extracellular protease activity mainly occurred at the onset of the stationary phase with a peak at 48 h in MM, which was consistent with previous results [ 25 ]. Moreover, the addition of gelatin led to a higher peak at the delayed time (72 h) in extracellular protease activity (Fig.…”

“…However, the production of extracellular proteases is considered to occur at the stationary growth phase and is extensively regulated in many species of Bacillus [25, 26]. For example, nitrogen and carbon sources have great impacts on the production of extracellular proteases [27].…”

“…B. pumilus BA06 has been isolated from proteinaceous soil and is demonstrated to be able to secrete extracellular proteases that exhibited great potential in leather processing [ 5 , 23 , 24 ]. However, the production of extracellular proteases is considered to occur at the stationary growth phase and is extensively regulated in many species of Bacillus [ 25 , 26 ]. For example, nitrogen and carbon sources have great impacts on the production of extracellular proteases [ 27 ].…”

Pasteurization of Lipid Emulsions with Supercritical CO2 and High Power Ultrasound

“…The emulsions were prepared in three stages: mixing, sonication and homogenization. The lipid phase was formed by soybean oil and egg phospholipid and Liu et al, 2015).…”

“…pumilus cells were sporulated following the methodology of , with modifications. A single colony of the bacteria was grown in nutrient broth (Scharlab, Spain) at 30ºC until the stationary phase was reached, according to bibliography (around 24 h) Liu et al, 2015). 100 µL of the culture with bacteria were poured on Plate Count Agar (Scharlab, Spain) enriched with MnSO4 (40 mg/L) and CaCl2 (100 mg/L) to enhance the sporulation and incubated at 30ºC for 5-6 days, a period of time in which spores were formed (confirmed with a Thoma counting chamber and an optical microscope).…”