The evolutionary origin of the mitochondrion: a nonsymbiotic model.
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Biomedical subjects
Publications and source records attributed to R A Raff.
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A cell-free protein-synthesizing system prepared from mouse sarcoma 180 was characterized by use of RNA from mengo virus and sea urchin egg. In the presence of exogenous mammalian transfer RNA, total sea urchin egg RNA and mengo RNA direct incorporation of [(3)H]leucine into acid-insoluble material. The system is extremely efficient in that a stimulation of 100-times over background can be obtained. Studies with formylmethionyl-transfer RNA, as well as with inhibitors of initiation, indicate that multiple initiation occurs; further, 85-90% of all chains made in vitro are subsequently released from ribosomes. An average translation time of 3.5 min was determined with messenger RNA of sea urchin egg, and product analysis indicates that high-molecular-weight products (greater than 50,000 molecular weight) are being made in vitro. Sequences of sea urchin egg RNA containing poly(A) act as messenger RNA.
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Uridinediphospho-N-acetyl glucosamine is converted to one or more uridine-diphospho-4-keto-6-deoxy-2-acetamidohexose derivatives by an enzyme from Citrobacter freundii ATCC 10053. Borohydride reduction of reaction products followed by acid hydrolysis yielded several amino sugars. Two of these were identified as fucosamine and quinovosamine by chromatography and ninhydrin degradation.
Studies employing colchicine binding, precipitation with vinblastine sulfate, and acrylamide gel electrophoresis confirm earlier proposals that Arbacia punctulata and Lytechinus pictus eggs and embryos contain a store of microtubule proteins. Treatment of 150,000 g supernatants from sea urchin homogenates with vinblastine sulfate precipitates about 5% of the total soluble protein, and 75% of the colchicine-binding activity. Electrophoretic examination of the precipitate reveals two very prominent bands. These have migration rates identical to those of the A and B microtubule proteins of cilia. These proteins can be made radioactive at the 16 cell stage and at hatching by pulse labeling with tritiated amino acids. By labeling for 1 hr with leucine-(3)H in early cleavage, then culturing embryos in the presence of unlabeled leucine, removal of newly synthesized microtubule proteins from the soluble pool can be demonstrated. Incorporation of labeled amino acids into microtubule proteins is not affected by culturing embryos continuously in 20 microg/ml of actinomycin D. Microtubule proteins appear, therefore, to be synthesized on "maternal" messenger RNA. This provides the first protein encoded by stored or "masked" mRNA in sea urchin embryos to be identified.
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Phenol-soluble lipopolysaccharides were obtained from the interphase and phenol phase fractions of 44% aqueous phenol-extracted Citrobacter species. Upon detailed investigation of C. freundii 8090, the two lipopolysaccharide fractions were found to contain different amounts of lipid A, although qualitative composition was similar. Both contained lipid A, 2-keto-deoxyoctonic acid, heptose, phosphate, d-glucose, galactose, rhamnose, 2-acetamido-2 deoxy-d-glucose, 3-acetamido-3,6-dideoxy-d-glucose, O-acetyl, and trace amino acids. Partially purified phenol-phase lipopolysaccharide partitioned into the phenol-soluble phase when refractionated with 44% aqueous phenol, and was further found to be soluble in 88% phenol, 95% ethyl alcohol, and chloroform-methanol (2:1).
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