Huiting Luo,Peijun Li,Hongwei Zhang,Xinping Diao,Baohua Kong
The objective of this study was to clarify whether formation of nitrosylmyoglobin (MbFe II NO) by Lactobacillus fermentum AS1.1880 in meat is due to nitric oxide synthase (NOS) activity. Confocal laser scanning microscopy exhibited strong green fluorescence in the L . fermentum sample treated with a nitric oxide (NO)-specific probe, directly indicating that NO was produced. Furthermore, determination of NOS activity based on the presence of NO metabolites indicated the existence of NOS in L.fermentum . A NOS inhibitor, N G -nitro-L-arginine methyl ester, significantly inhibited the activity of NOS in L.fermentum ( P < 0.05). Futhermore, NOS protein was detected in L.fermentum by Western blot analysis. L-arginine addition largely increased the NOS activity of L.fermentum ( P < 0.05). In meat batters , the redness of a sample inoculated with L.fermentum was higher than that of the control and colour was significantly improved with the addition of L-arginine ( P < 0.05), indicating that more MbFe II NO was formed.
Xu Chenglong,Lin Wenxi,Chen Yunni,Gao Boliang,Zhang Zhibin,Zhu Du
Perylenequinones (PQs) are a class of natural polyketides used as photodynamic therapeutics. Heat stress (HS) is an important environmental factor affecting secondary metabolism of fungi. This study investigated the effects of HS treatment on PQs biosynthesis of Shiraia sp. Slf14(w) and the underlying molecular mechanism. After the optimization of HS treatment conditions, the total PQs amount reached 577 ± 34.56 mg/L, which was 20.89-fold improvement over the control. Also, HS treatment stimulated the formation of intracellular nitric oxide (NO). Genome-wide analysis of Shiraia sp. Slf14(w) revealed iNOSL and cNOSL encoding inducible and constitutive NOS-like proteins (iNOSL and cNOSL), respectively. Cloned iNOSL in Escherichia coli BL21 showed higher nitric oxide synthase (NOS) activity than cNOSL, and the expression level of iNOSL under HS treatment was observably higher than that of cNOSL , suggesting that iNOSL is more responsible for NO production in the HS-treated strain Slf14(w) and may play an important role in regulating PQs biosynthesis. Moreover, the putative biosynthetic gene clusters for PQs and genes encoding iNOSL and nitrate reductase (NR) in the HS-treated strain Slf14(w) were obviously upregulated. PQs biosynthesis and efflux stimulated by HS treatment were significantly inhibited upon the addition of NO scavenger, NOS inhibitor, and NR inhibitor, indicating that HS-induced NO, as a signaling molecule, triggered promoted PQs biosynthesis and efflux. Our results provide an effective strategy for PQs production and contribute to the understanding of heat shock signal transduction studies of other fungi.
Key points
• PQs titer of Shiraia sp. Slf14(w) was significantly enhanced by HS treatment . • HS-induced NO was first reported to participate in PQs biosynthetic regulation . • Novel inducible and constitutive NOS-like proteins (iNOSL and cNOSL) were obtained and their NOS activities were determined .
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