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1.
乳酸乳酸球菌AL2产生的乳链菌肽的提纯和性质   总被引:12,自引:2,他引:10  
用NaCl饱和的乳酸乳酸球菌(Lactococcus lactis subsp. Lactis)AL2发酵液经正丙醇提取和CM-Sephadex C-25柱层析,得到聚丙烯酰胺凝胶电泳纯的乳链菌肽组分,比活力从24427IU/mg提高到39865IU/mg,活力回收为41.7%。Α—胰凝乳蛋白酶可使乳链菌肽丧失活性;在低pH条件下,乳链菌肽对热较稳定;对许多革兰氏阳性菌有强烈抑制作用,而对革兰氏阴性菌、酵母菌和霉菌没有作用。  相似文献   

2.
用本实验室获得的乳链菌肽高产菌株乳酸乳球菌AL2总DNA为供体,以λ EMBL3为载体,构建了该菌株的基因文库,共获得3900多个噬斑。通过Southern杂交、PCR扩增及DNA序列测定,证实从该文库中筛选到一个含有完整乳链菌肽生物合成基因簇的重组噬菌体λHJ\|3。  相似文献   

3.
一个含有乳链菌肽抗性基因的乳酸乳球菌质粒pTS50的鉴定   总被引:5,自引:0,他引:5  
在添加乳链菌肽、乳糖及溴甲酚紫的M1 7选择培养基上 ,从 1 97个新鲜牛奶样品中筛选到 3株乳链菌肽抗性菌株 ,PCR扩增证实它们都含有乳链菌肽抗性基因。菌种生理生化特性鉴定及特异性 1 6SrDNAPCR扩增产物的序列测定结果表明这 3株菌都属于乳酸乳球菌乳酸亚种。质粒转化实验发现乳酸乳球菌乳酸亚种TS 1 640中的乳链菌肽抗性基因位于一个约47kb的大质粒pTS50上。BamHI、EcoRI、HindⅢ、NcoI、PstⅠ酶切分析和Southern杂交 ,进一步将乳链菌肽抗性基因定位于pTS50的一个约 1 9kbEcoRI酶切片段中  相似文献   

4.
乳链菌肽前体基因(nisZ)在乳酸乳球菌中的克隆和表达   总被引:8,自引:1,他引:7  
用PCR技术从克隆有完整乳链菌肽生物合成基因簇(来自于乳链菌肽高产菌株L.lactis AL2)的重组噬菌体λHJ-3中扩增了编码乳链菌肽的前体基因,与pMG36e连接得到重组质粒pHJ201,用电击转化法将pHJ201转化到L.lactis NZ9800中,经活性测定和Tricine-SDS-PAGE电泳证实乳链菌肽前体基因获得了功能表达。DNA序列分析表明乳链菌肽高产菌株L.lactis AL2产生的是NisinZ。发现pHJ201d L.lactis NZ9800 中有良好的稳定性。  相似文献   

5.
乳链菌肽的分离纯化和部分生物学性质   总被引:1,自引:0,他引:1  
用乳酸链球菌SM526进行乳链菌肽的发酵生产,产量为40~50mg/l.经中空纤维超滤器超滤,非极性大孔吸附树脂XAD-2层析,CM-SephadexC-25层析和SephadexG-50层析纯化了该肽。SDS-PAGE表明达均一,RP-HPLC表明其纯度不低于95%。SDS-PAGE测其Mr约为3600,用IEF测其等电点为9.5.酸性条件下稳定且抗热;对胰蛋白酶、胃蛋白酶和木瓜蛋白酶不敏感,但对α-胰凝乳酶和蛋白酶K敏感。乳链菌肽对多种革兰氏阳性菌有强烈的抑制作用;以枯草杆菌和金黄色葡萄球菌为指示菌,其作用方式是杀菌。  相似文献   

6.
乳链菌肽产生菌的定向筛选及发酵产物的鉴定   总被引:9,自引:0,他引:9  
利用乳链菌肽产生菌中nip^+nis^rsuc^+紧密连锁的原理,在添加乳链菌肽、蔗糖及溴甲酚紫的选择培养基上,从牛奶样品中定向筛选乳链菌肽产生菌。对筛选到的L. lactis1409菌株发酵产物的分析鉴定结果揭示:该产物对多种革兰氏阳性菌有强烈抑制作用,而对革兰氏阴性菌、酵母菌和霉菌无效,在pH值低的条件下对热稳定,对α—胰凝乳蛋白酶敏感,具有生物活性的蛋白质的分子量与乳链菌肽相当,而1409菌株的质粒分布与L. laclisATCC11454和L. lactis 7962不同,说明筛选到的L. laclis 1409菌株确是一株新的乳链菌肽产生菌。  相似文献   

7.
乳酸抗性酵母的筛选及其生长特性的研究   总被引:2,自引:0,他引:2  
以酿酒酵母 (saccharomycesceevisiae)单倍体YNN -2 7(αtep ura )为亲株 ,在含有 4 %乳酸的梯度平板上直接进行紫外线诱变处理 ,筛选到突变株YNN -2 7-2 4。通过对该突变株乳酸抗性产生原因分析、在含有不同浓度的乳酸和潮霉素B(hygromycinB)的YPDL和YPDLH培养基中的重复特性的研究发现 ,该突变株对乳酸和潮霉素B产生的抗性 ,不是因对环境条件的适应而产生 ,而是由基因突变所引起。与突变株YNN2 7-2 4相比 ,乳酸对亲株生长的影响在于延长了其生长的延迟期 ,而其生长速率没有发生改变。用Mini-photo 51 8测定供试菌株在生长过程中的吸光度 ( 660nm)以研究酵母菌的生长特性 ,是一种行之有效的方法 ,具有较高的灵敏度和较好的再现性。  相似文献   

8.
20 0 0年 10月 2 5日 ,国家“九五”攻关项目—乳链菌肽的工业化生产在中国科学院微生物研究所通过专家鉴定。至此 ,乳链菌肽—这个凝聚着科研人员十余年心血的天然食品防腐剂终于从实验室走进市场。乳链菌肽 (Nisin)亦称之为乳酸链球菌素或乳酸链球菌肽 ,是乳酸链球菌 (现定名为乳酸乳球菌 )产生的分子量约为 350 0 D的一种小肽。对引起食品腐败的许多革兰氏阳性菌具有强烈抑杀作用 ,已被世界许多国家和地区广泛用作乳制品、植物蛋白食品、罐头食品、肉制品等食品的防腐保鲜。我国对乳链菌肽的研究始于 1989年 ,中国科学院微生物研究所的…  相似文献   

9.
目的:通过基因工程方法提高HPr蛋白编码基因ptsH在乳链菌肽(nisin)高产野生乳酸乳球菌株N8中的表达,揭示ptsH基因与乳酸乳球菌乳链菌肽耐受性等相关生物学功能的关系。方法:构建ptsH过表达质粒pLEV16-ptsH并转化至N8,使其ptsH基因过量表达,进而对比分析ptsH过表达菌株与野生菌株在生长曲线、乳链菌肽耐受性、效价、Biolog等方面的差异。结果:N8-ptsH过表达菌株与N8菌株在菌落形态、大小、表面湿滑程度及生长曲线等方面没有明显差异;ptsH基因过表达使N8菌株的乳链菌肽耐受性提高了8.3%,2个乳链菌肽耐受性相关基因nisI和nisF的表达量分别提高了15.45倍和近45倍;ptsH基因过表达略微减缓了N8菌株中乳链菌肽的产生,但乳链菌肽的最终产量略有提高;ptsH基因过表达菌株中PTS系统糖苷和磷酸化糖类的利用率比原始菌株显著提高。结论:ptsH基因主要与乳酸乳球菌的乳链菌肽耐受性有关。  相似文献   

10.
质子自杀法选育克雷伯氏菌产乳酸突变株   总被引:1,自引:1,他引:0  
以产酸克雷伯氏菌(Klebsiella oxytoca) M5al为出发菌株, 经亚硝基胍诱变处理, 运用质子自杀法选育, 从含0.17 mol/L NaBr-NaBrO3的初筛平板上选出44个具有稳定遗传性的单菌落, 然后结合培养基优化后的摇瓶发酵复筛, 获得3个产乳酸突变株, 其乳酸脱氢酶活性分别为出发菌株的50.6%、58.8%、61.3%。对其中乳酸脱氢酶活性最低的菌株在5 L自动发酵罐上进行批式发酵, 结果显示: 突变株乳酸产量大幅降低, 而乙酸、1,3-丙二醇的产量则显著增加。  相似文献   

11.
SUMMARY: Spores of two strains of Bacillus stearothermophilus were heated for various times both in the absence and presence of nisin and in the latter case both with and without subsequent destruction of the nisin with trypsin. The results obtained on enumeration of the survivors indicated that the reduction of heat resistance which was observed was apparent rather than real and was due to the adsorption of nisin on the spores. When this was removed by means of trypsin the heat resistance was restored almost to normal.  相似文献   

12.
SUMMARY: The action of various concentrations of nisin on the development of cells and spores from untreated and heated suspensions of Clostridium welchii added to gelatine has been investigated, using a tube colony count technique. The development of vegetative cells was prevented by the presence of 2 Reading Units (R.U.) of nisin/ml of the final culture medium, although the effective concentration may have been 6 R.U./ml. Approximately 40 R.U./ml prevented colony formation from spores. Gelatine containing nisin was dried. A considerable proportion of the activity of the antibiotic was still present after storage for 5 weeks.  相似文献   

13.
固定化乳酸乳球菌连续生产Nisin的研究   总被引:6,自引:0,他引:6  
以海藻酸钙为材料 ,固定乳酸乳球菌 (Lactococcuslactissubsp .lactis)SM5 2 6 ,研究不同条件对Nisin合成的影响。结果表明 ,利用 2 %海藻酸钠在 1 0mmol LCaCl2 条件下 ,得到的固定化细胞颗粒稳定性较好 ,可维持 90h无破裂 ;在发酵过程中SYS3培养基中的无机盐成分尤其磷酸盐对固定化颗粒有破坏作用 ;用mSYS3培养基代替SYS3 ,通过 72h三批次循环的半连续培养 ,Nisin活性为 85 0IU mL ,无明显的细胞渗漏现象。连续化生产 70h ,Nisin活性达 1 1 5 0IU mL ,相当于游离细胞的发酵水平。  相似文献   

14.
A SIMPLE PROCEDURE TO DETECT NISIN IN CHEESE   总被引:4,自引:1,他引:3  
SUMMARY: A suitable amount of agar medium inoculated with a test organism sensitive to nisin ( Lactobacillus lactis or Streptococcus cremoris ) is poured on a cheese cylinder placed in a Petri dish. After a convenient incubation time the presence of a clear zone around the cheese cylinder will indicate the presence of nisin.
The method is applicable both to natural and processed cheese and can also be used for other antibiotics that may be present in cheese.  相似文献   

15.
A nisin-resistant (NISr) variant of Listeria monocytogenes Scott A was isolated by stepwise exposure to increasing concentrations of nisin in brain heart infusion (BHI) broth. The NISr strain was about 12 times more resistant to nisin than was the wild-type (WT) strain. Accordingly, higher nisin concentrations were required to dissipate both components of the proton motive force in the NISr strain than in the WT strain. Comparison of the membrane fatty acyl composition of the sensitive strain with that of its NISr derivative revealed no significant differences. From phospholipid head group composition analysis and phospholipid biosynthesis measurements during growth in the absence and presence of nisin, it could be inferred that the NISr strain produces relatively more phosphatidylglycerol (PG) and less diphosphatidylglycerol (DPG) than the parent strain does. Monolayer studies with pure lipid extracts from both strains showed that nisin interacted more efficiently with lipids derived from the WT strain than with those derived from the NISr strain, reflecting qualitative differences in nisin sensitivity. Involvement of the cell wall in acquisition of nisin resistance was excluded, since the WT and NISr strains showed a comparable sensitivity to lysozyme. Recently, it has been demonstrated that nisin penetrates more deeply into lipid monolayers of DPG than those of other lipids including PG, phosphatidylcholine, phosphatidylethanolamine, monogalactosyldiacylglycerol, and digalactosyldiacylglycerol (R.A. Demel, T. Peelen, R.J. Siezen, B. de Kruijff, and O.P. Kuipers, Eur. J.Biochem. 235:267-274, 1996). Collectively, the mechanism of nisin resistance in this L. monocytogenes NISr strain is attributed to a reduction in the DPG content of the cytoplasmic membrane.  相似文献   

16.
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