Disruption of the external sheath of Streptomyces granaticolor aerial spores and subsequent cultivation in a rich medium result in a synchronous germination. This method was used to analyze RNA and protein patterns during the germination. The germination process took place through a sequence of time-ordered events. RNA and protein synthesis started during the first 5 min and net DNA syntesis at 60-70 rnin of germination. Within the first 10 min of germination, synthesis of RNA was not sensitive to the inhibitory effect of rifamycin. During this period rRNA and other species including 4 -5-S RNA were synthesized. Dormant spores contained populations of ribosomes or ribosomal precursors that were structurally and functionally defective. The ribosomal particles bound a sporulation pigment(s) of the melanine type. The ribosomal proteins complexed to the pigments formed insoluble aggregates which were easily removed from the ribosomes by one wash with 1 M NH4Cl. During the first 10 rnin of germination, pigment(s) were liberated from the complexes with the ribosomes and protein extracts of the washed ribosomes had essentially the same pattern as the extracts of ribosomes of vegetative cells. These structural alterations were accompanied by enhancement of the ribosome activities in polypeptide synthesis in vivo and in vitro. When the spores were incubated with a I4C-labelled amino acid mixture in the presence of rifamycin, only three proteins (GSI, GL1 and GS9) were identified to be radiolabelled in the extracts from the washed ribosomes. These experiments indicate that liberation of the sporulation pigment(s) from the complexes with ribosomal proteins and assembly of de novo synthesized proteins and proteins from a preexisting pool in the spore are involved in the reactivation of the ribosomes of dormant spores of S. granaticolor.The control of synthesis of specific proteins and their organization into complex biological structures is the central problem of cell differentiation. Germination of aerial spores of various Streptomyces strains is an ordered developmental process, whereby a dormant spore is converted into a growing vegetative cell [I -31. These events are accompanied by sequential reactivation of transcription, translation and DNA replication systems [4, 51.Most of our present knowledge about transcriptional and translational control and how they operate derives mainly from genetic and biochemical studies of gene expression in Escherichia coli and Bacillus subtilis.Reports from several laboratories indicated that the differentiation in microorganisms is regulated both at the level of transcription [6-81 and translation [9, 101. Studies on the protein-synthesizing system of vegetative cells and dormant spores revealed differences in many properties including alterations of ribosomal proteins [I 1 -141, transfer RNA [15-171 and activity or specificity at the translation of synthetic and natural messenger RNA [18, 191. In order to understand the mechanisms involved in the control of activity of the translati...
The rates of RNA, protein and DNA synthesis were estimated in synchronously germinating spores of Streptomyces granaticolor. Rapid uptake of labelled precursors of RNA and proteins was observed after 20 s. The germination process took place through a sequence of time-ordered events. RNA synthesis started after 3 min of germination, protein synthesis began at 4 min and net DNA synthesis at 60-70 min of germination. A characteristic feature of germination was the biphasic pattern in the rate of RNA and protein synthesis. Spores of Streptomyces granaticolor were sensitive to actinomycin D, rifampicin and chloramphenicol even at the start of germination. Protein synthesis during germination was dependent on new mRNA synthesis and was independent during the first 60-70 min on replication of the spore genome.
The ability of EF‐Tu to aggregate spontaneously was employed for the purification of homogeneous EF‐Tu · GDP from Streptomyces aureofaciens. The formation of filamentous structures in the aggregated EF‐Tu was demonstrated in a light microscope. The purified factor, with a specific activity of 19 100 ± 1000 units/mg in [3H]GDP exchange, was shown to be active in the translation of poly(U).
Aggregated EF‐Tu · GDP exhibited almost eight‐times lower GDP‐exchange capacity at 2°C than at 30°C. This suggests that GDP‐binding sites are not freely accessible at lower temperatures in the aggregated factor, in contrast to Escherichia coli polymerized EF‐Tu. Turbidimetric assays revealed that the solubilization of diluted aggregated S. aureofaciens EF‐Tu is strongly dependent on temperature and causes an increase in the number of accessible GDP‐binding sites.
Ribosomes from cells of Streptomyces aureofaciens producing tetracycline antibiotics (Tc‐ribosomes) differ in electrophoretic mobility of ribosomal proteins S2, S10 and L19 from those of the same strain, where the production of tetracyclines was suppressed by changed cultivation conditions (C‐ribosomes). Purified tight vacant couples C‐ and Tc‐ribosomes are equally active in the translation of poly(U). Both types of S. aureofaciens ribosomes are more sensitive to tetracycline and chlortetracycline than ribosomes of Escherichia coli in the Phe‐tRNA binding and the translation of poly(U).
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