产细菌素乳酸菌的筛选及其发酵特性

从传统发酵制品中分离出60株乳酸菌,在排除有机酸及过氧化氢的干扰后,从中筛选出抑菌活性较高的菌株P-319,经常规生理生化实验,鉴定该菌株为短乳杆菌。其发酵上清液经胰蛋白酶和木瓜蛋白酶处理后抑菌活性丧失;对其抑菌谱的测定表明,其能抑制革兰氏阳性菌和革兰氏阴性菌的生长,但对霉菌和酵母菌没有任何抑制作用。产细菌素的发酵过程测定结果表明,该菌株在MRS培养基中30℃培养24 h时,抑菌活性达到最高。     

High‐level extracellular production of penicillin acylase by genetic engineering of Escherichia coli

The extracellular production of penicillin acylase (PAC) in genetically engineered Escherichia coli by coexpression of the brp gene encoding bacteriocin release protein (BRP) and the pac gene was demonstrated. Cell physiology was affected while PAC was released into the medium, depending on the strategy for brp expression. The performance for the production and release of PAC was optimized by taking several culture parameters, including host, inducer (mitomycin C) concentration, and induction timing for brp expression, into consideration. The effect of PAC release on inclusion body formation was also investigated. It was observed that the amount of inclusion bodies was significantly affected by brp expression. A reason for the limitation of PAC production and a strategy for resolving this problem are proposed. © 2001 Society of Chemical Industry     

嗜酸乳杆菌WS所产细菌素生物学特性的研究

LactobacliiusacidophilusWS在MRS培养基中经过37℃培养得到的细菌素经受70℃、121℃处理20min后,活性几乎不变,对以金黄色葡萄球菌为代表的革兰氏阳性菌有明显的抑制作用,而对大肠杆菌为代表的革兰氏阴性菌的抑制作用不明显,说明该细菌素是一种广谱抗菌的细菌素。细菌素对木瓜蛋白酶(papain)、蛋白酶K(proteinaseK)以及胰蛋白酶(trypsin)敏感,而对中性蛋白酶(subtilisin)和胃蛋白酶(pepsin)的敏感性差。在一定的范围内,细菌素的抑菌活性和环境的酸度呈正相关。     

Effect of Exogenous Factors on Bacteriocin Production from Lactobacillus paracasei J23 by Using a Resting Cell System

A resting cell system was developed for bacteriocin Lac-B23 production from Lactobacillus paracasei J23. The resting cell medium contained (g/L): Glucose 20, Sodium acetate 5.0, MnSO₄ 0.25 MgSO₄ 0.5, Ammoniumhydrogencitrate 1.0, KH₂PO₄ 1.0. The resting cell incubation time and temperature were 20 h and 37 °C and the effects of exogenous factors, including amino acids, glycerol, pyruvic acid, and α-ketoglutaric acid were investigated. Cys and Gly could stimulate the production of bacteriocin, while no stimulus effect was observed for Glu, Tyr and Ala. Glycerol and pyruvic acid increased bacteriocin production and the optimum concentrations were 1% and 30 g/L, respectively. Bacteriocin could act as an inducer of its own biosynthesis. These findings are of importance for the further study of bacteriocin biosynthesis regulation and for the improvement of bacteriocin production yields.     

Effect of high-pressure treatment and a bacteriocin-producing lactic culture on the proteolysis, texture, and taste of Hispánico cheese

The effects of high-pressure treatment, by itself or in combination with a bacteriocin-producing culture added to milk, on the proteolysis, texture, and taste of Hispánico cheese were investigated. Two vats of cheese were manufactured from a mixture of cow and ewe milk. Milk in one vat was inoculated with 0.5% Lactococcus lactis ssp. lactis INIA 415, a nisin Z and lacticin 481 producer; 0.5% L. lactis ssp. lactis INIA 415-2, a bacteriocin-nonproducing mutant; and 2% of a commercial Streptococcus thermophilus culture. Milk in the other vat was inoculated with 1% L. lactis ssp. lactis INIA 415-2 and 2% S. thermophilus culture. After ripening for 15 d at 12°C, half of the cheeses from each vat were treated at 400 MPa for 5 min at 10°C. Ripening of high-pressure-treated and untreated cheeses continued at 12°C until d 50. High-pressure treatment of cheese made from milk without the bacteriocin producer accelerated casein degradation and increased the free AA content, but it did not significantly influence the taste quality or taste intensity of the cheese. Addition of the bacteriocin producer to milk lowered the ratio of hydrophobic peptides to hydrophilic peptides, increased the free AA content, and enhanced the taste intensity. The combination of milk inoculation with the bacteriocin producer and high-pressure treatment of the cheese resulted in higher levels of both hydrophobic and hydrophilic peptides but had no significant effect on the free AA content, taste quality, or taste intensity.     

Experimentally induced intramammary infection with multiple strains of Streptococcus uberis

The effect of infusing a mixture of 5 Streptococcus uberis strains into mammary quarters of 10 lactating cows was investigated. All 5 strains, which included 2 originally isolated from the dairy environment and 3 from clinical cases of mastitis, were capable of establishing an intramammary infection when infused individually. However, when the 5 strains were infused together, a single strain predominated in 7 out of 10 quarters. One strain in particular prevailed in 4 mammary quarters and was also found to inhibit the growth of the other 4 strains with deferred antagonism on esculin blood agar. The genes required for the production of bacteriocins nisin U and uberolysin were identified in this strain, whereas the other 4 strains contained only uberolysin genes. Direct competition may have occurred between strains within the mammary gland but competition was not apparent when cultured together in UHT milk, where no strain predominated. Although the mechanism is unknown, these results imply that a selection process can occur within the mammary gland, leading to a single strain that is detected upon diagnosis of mastitis.     

Structure,Biosynthesis, and Biological Activity of Succinylated Forms of Bacteriocin BacSp222

BacSp222 is a multifunctional peptide produced by Staphylococcus pseudintermedius 222. This 50-amino acid long peptide belongs to subclass IId of bacteriocins and forms a four-helix bundle molecule. In addition to bactericidal functions, BacSp222 possesses also features of a virulence factor, manifested in immunomodulatory and cytotoxic activities toward eukaryotic cells. In the present study, we demonstrate that BacSp222 is produced in several post-translationally modified forms, succinylated at the ε-amino group of lysine residues. Such modifications have not been previously described for any bacteriocins. NMR and circular dichroism spectroscopy studies have shown that the modifications do not alter the spatial structure of the peptide. At the same time, succinylation significantly diminishes its bactericidal and cytotoxic potential. We demonstrate that the modification of the bacteriocin is an effect of non-enzymatic reaction with a highly reactive intracellular metabolite, i.e., succinyl-coenzyme A. The production of succinylated forms of the bacteriocin depends on environmental factors and on the access of bacteria to nutrients. Our study indicates that the production of succinylated forms of bacteriocin occurs in response to the changing environment, protects producer cells against the autotoxicity of the excreted peptide, and limits the pathogenicity of the strain.