"Acidic pH-sensitive mutants": acid resistance in acetic acid bacteria

1)采用亚硝基胍处理，再用常规复型法从葡萄糖酸杆菌IFO 3267中分离到15株酸性pH敏感突变株。分离的菌株显示出溶解的性质<CaCO_3>，不存在氧化乳酸和乙酸为二氧化碳和水以及存在泛醌-10。菌株的无细胞提取物分别具有膜结合葡萄糖脱氢酶、5-酮果糖还原酶、甘露醇脱氢酶等酶活性。2）所有突变菌株在pH 4.0时均为阴性生长。在所分离的突变菌株中，PSM-51、PSM-64、PSM-104、PSM-110、PSM-120和PSM-121这6株菌株对酸性和碱性pH特别敏感。它们在pH 6.0时表现出最大生长。然而，在中性pH的两侧，即，pH5.0和pH7.0时，其生长逐渐减弱 ...更多信息 分析了所有突变株的脂质组成。3株突变株PSM-51、PSM-120和PSM-121的磷脂组成与出发菌株相比存在差异。首先，PSM-51的磷脂酰乙醇胺含量从2倍增加到3倍，磷脂酰胆碱含量从25%降低到30%。其次，突变株PSM-120和PSM-121的磷脂含量比出发菌株提高了60%。4）利用PSM-51的完整细胞和含[甲基- &lt;^（14）C&gt; ]蛋氨酸的无细胞提取物，分析了PSM-51由磷脂酰乙醇胺合成磷脂酰胆碱的生物合成途径。突变株在完整细胞中的活性为60%，在无细胞提取物中的活性与亲本菌株相同。5）用放射性弱有机酸5，5-二甲基[2-（14）C&gt; ]恶唑烷-2，4-二酮跨细胞膜测定突变株和亲本菌株的细胞内和细胞外pH。在亲本菌株中，细胞内pH不受细胞外pH改变的影响，即，从pH 3.0到pH 7.0：细胞内pH几乎恒定地保持在6.0-6.5的pH范围内。另一方面，当在pH 4.0下孵育时，所有突变株的细胞内pH降低至pH 5.9以下。在胞外pH 3.0时，不能测量突变株的胞内pH。亲本菌株的细胞内pH保持在pH 6.03。少

1) Fifteen strains of acidic pH-sensitive mutants were isolated from Gluconobacter cerinus IFO 3267 by treatment of nitrosoguanidine, followed by the conventional replica method. The isolated strains showed the properties of the dissolution of <CaCO_3> , the absence of oxidizing lactate and acetate to carbon dioxide and water and the presence of ubiquinone-10. The cell-free extracts of the strains had the respective enzyme activities of membrane-bound glucose dehydrogenase, 5-ketofructose reductase, mambrane-bound mannitol dehydrogenase and so on.2) The growth of all the mutant strains was negative at pH 4.0. However, the better growth was observed at and near the neutral pH. Of the isolated mutant strains, the six strains of PSM-51, PSM-64, PSM-104, PSM-110, PSM-120 and PSM-121 were especially sensitive to the acid and alkaline pHs. They showed maximal growth at pH 6.0. However, in the both sides of the neutral.pH, i.e., pH 5.0 and pH 7.0, their growth gradually decreased.3) The membr … More ane lipid composition of all the mutant strains were analyzed. Three mutant strains, PSM-51, PSM-120 and PSM-121 were different in phospholipid composition compared with the parent strain. Firstly, PSM-51 was increased in the content of phosphatidylethanolamine from two to three times and decreased in that of phosphatidylcholine from 25 to 30%. Secondly, other two mutant strains, PSM-120 and PSM-121 had 60% phospholipid content compared with the parent strain.4) The biosynthetic pathway of phosphatidylcholine from phosphatidylethanolamine was analyzed by using the intact cells and the cell-free extract of PSM-51 with [methyl- <^(14)C> ]methionine. The activities of the mutant strain was 60% in the intact cells and the same in the cell-free extract compared with the parent strain.5) The intracellular and extracellular pHs of the mutant and the parent strains were measured across the cell membrane with a radioactive weak organic acid, 5,5-dimethyl[2- <^(14)C> ]oxazolidine-2,4-dione. In the parent strain, the intracellular pH was not affected by the alteration of the extracellular pH, i.e., from pH 3.0 to pH 7.0: the intracellular pH was almost constantly retained in the pH range of 6.0-6.5. On the other hand, the intracellular pH of all the mutant strains decreased below pH 5.9 when incubated at pH 4.0. At the extracellular pH 3.0, the intracellular pHs of the mutant strains were not able to be measured. The intracellular pH of the parent strain was retained at pH 6.03. Less