Mechanism of heat resistance of bacterial spores

为了阐明细菌芽胞的耐热性，进行了以下实验。枯草芽孢杆菌休眠孢子在4ﾟ℃、静水压力1000~5000 kg/cm~2范围内处理1h后仍能存活，对枯草杆菌休眠孢子的影响不大。在静水压力相同的条件下，37ﾟC和60ﾟC处理的休眠孢子绝大多数被杀死，所观察到的现象类似于L-丙氨酸的生理萌发现象。休眠孢子中非活性葡萄糖脱氢酶的纯化。葡萄糖脱氢酶被认为在萌发后立即在能量产生中起重要作用，我们估计该酶在休眠孢子中以失活形式存在，该酶由两个相对分子质量为30K的亚基组成，并在萌发过程中成为具有四个亚基的活性形式。因此，利用抗192-207肽的抗体和巨大芽孢杆菌的酶活力，尝试从枯草杆菌芽胞中分离纯化失活型枯草杆菌孢子酶。后一种抗体似乎是可用的。二吡啶甲酸(DPA)与纯化的失活酶的结合将与孢子的耐热性有关。休眠孢子中DPA结合大分子物质的检测和纯化虽然细菌孢子中含有丰富的DPA，但其作用尚不清楚。因此，我们一直在尝试利用抗DPA的抗体来检测枯草杆菌休眠孢子中的一些DPA结合的大分子物质。检测到两种HW分别为440K和230K的物质，其纯化正在进行中。将研究DPA在耐热性中的作用。

The following experiments were carried out for the purpose of clarification of the heat resistance of bacterial spores.1. Death of bacterial spores by hydrostatic pressure at various temperatures.The dormant spores of Bacillus subtilis treated at 4ﾟC for 1 h with hydrostatic pressures between 1, 000 and 5, 000 kg/cm^2 remained alive without being affected very much. On the other hand, a great majority of dormant spores treated at 37ﾟC and 60ﾟC under the same conditions of hydrostatic pressure were killed and the phenomena observed resembled those of physiological germination by L-alanine.2. Purification of inactive glucose dehydrogenase from dormant spores.Glucose dehydrogenase is considered to play an important role in energy production immediately after germination and we have estimated that the enzyme is present in dormant spores as an inactive form consisting of two subunits with M. W. of 30K and becomes an active form having four subunits during germination. Therefore, purification of the inactive form of the enzyme from B. subtilis spores has been tried using the antibodies raised against the 192-207 peptide and the active fore of the enzyme from Bacillus megaterium. The latter antibody appeared to be available. The binding of dipicolinic acid (DPA) to the purified inactive enzyme would be examined in connection with heat resistance of spores.3. Detection and purification of DPA-binding macromoleculer substances in dormant spores.Although DPA is contained abundantly in bacterial spores, its role remains unclear. Therefore, we have been trying to detect some DPA-binding macromolecular substances in dormant spores of B. subtilis using the antibody against DPA. Two substances with H. W. of 440K and 230K were detected and their purification is in progress. The role of DPA in heat resistance would be examined.