MUTANTS OF ESCHERICHIA-COLI DEFECTIVE IN MEMBRANE PHOSPHOLIPID SYNTHESIS - MACROMOLECULAR-SYNTHESIS IN AN SN-GLYCEROL 3-PHOSPHATE ACYLTRANSFERASE KM MUTANT
MUTANTS OF ESCHERICHIA-COLI DEFECTIVE IN MEMBRANE PHOSPHOLIPID SYNTHESIS - MACROMOLECULAR-SYNTHESIS IN AN SN-GLYCEROL 3-PHOSPHATE ACYLTRANSFERASE KM MUTANT
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DOI：
10.1128/jb.117.3.1065-1076.1974
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发表时间：
1974-01-01
影响因子：
3.2
通讯作者：
BELL, RM
中科院分区：
文献类型：
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作者：
BELL, RM
从一株不能有氧分解代谢3-磷酸甘油（G3 P）的大肠杆菌中筛选出G3 P营养缺陷型菌株。营养缺陷型由生物合成G3 P脱氢酶（EC 1.1.1.8）的损失或由有缺陷的膜G3 P酰基转移酶引起。在突变体的膜制备物中，G3 P酰基转移酶的表观Km比亲本的高11- 14倍（从约90 μm到1，000 - 1，250 μm）。从G3 P脱氢酶突变体的回复突变体制备的所有提取物均显示G3 P脱氢酶活性，但大多数提取物的含量低于野生型水平的10%。从酰基转移酶突变体的回复突变体的膜制剂有appartKm的G3 P类似的父母。已经衍生出菌株，其中在葡萄糖的存在下可以用甘油满足G3 P需求，这可能是因为甘油激酶对1，6-二磷酸果糖的抑制脱敏。对G3 P酰基转移酶Km突变体的生长和大分子合成的研究表明，甘油剥夺后，净磷脂合成立即停止，生长持续约一倍，净核糖核酸（RNA）、脱氧核糖核酸（DNA）和蛋白质几乎加倍，与生长曲线平行;通过用32 P-磷酸、14 C-乙酸或3 H-丝氨酸标记细胞评估的磷脂合成速率降低大于90%; RNA和DNA的合成速率随着细胞生长而增加，然后随着细胞停止生长而降低; 2蛋白质的合成速率没有显示出增加，并且当细胞停止生长时比RNA和DNA的合成速率下降得更慢。甘油剥夺后，细胞保持并获得合成磷脂的能力。这些数据表明，净磷脂合成是不需要继续大分子合成约一倍，这些过程的速率不耦合在这段时间内。
sn-Glycerol 3-phosphate (G3P) auxotrophs ofEscherichia colihave been selected from a strain which cannot aerobically catabolize G3P. The auxotrophy resulted from loss of the biosynthetic G3P dehydrogenase (EC 1.1.1.8) or from a defective membranous G3P acyltransferase. The apparentKmof the acyltransferase for G3P was 11- to 14-fold higher (from about 90 μm to 1,000 to 1,250 μm) in membrane preparations from the mutants than those of the parent. All extracts prepared from revertants of the G3P dehydrogenase mutants showed G3P dehydrogenase activity, but most contained less than 10% of the wild-type level. Membrane preparations from revertants of the acyltransferase mutants had apparentKm's for G3P similar to that of the parent. Strains have been derived in which the G3P requirement can be satisfied with glycerol in the presence of glucose, presumably because the glycerol kinase was desensitized to inhibition by fructose 1,6-diphosphate. Investigations on the growth and macromolecular synthesis in a G3P acyltransferaseKmmutant revealed that upon glycerol deprivation, net phospholipid synthesis stopped immediately; growth continued for about one doubling; net ribonucleic acid (RNA), deoxyribonucleic acid (DNA), and protein nearly doubled paralleling the growth curve; the rate of phospholipid synthesis assessed by labeling cells with32P-phosphate,14C-acetate, or3H-serine was reduced greater than 90%; the rates of RNA and DNA synthesis increased as the cells grew and then decreased as the cells stopped growing; the rate of protein synthesis showed no increase and declined more slowly than the rates of RNA and DNA synthesis when the cells stopped growing. The cells retained and gained in the capacity to synthesize phospholipids upon glycerol deprivation. These data indicate that net phospholipid synthesis is not required for continued macromolecular synthesis for about one doubling, and that the rates of these processes are not coupled during this time period.