Actinomycins sequence

The NMR experiments 55 are obtained from actinomycin D in order to check the amino acid sequence, to assign proton-proton and some carbon-proton connectivities, and to deduce informations concerning proton distances and the spatial structure of both cyclopentapeptide lactone rings. Conditions CDCI3, 10 mg per 0.3 ml, 25 °C, 500 MHz H), 125 MHz ( C). (a) HH COSY plot ... 

Many NRPs such as cyclosporin, complestatin, actinomycin, and chondramide contain N-methyl amides. M-Methyl transferase (N-MT) domains utilize S-adenosylmethionine (SAM) as a cofactor to catalyze the transfer of the methyl group from SAM to the a-amine of an aminoacyl-S-PCP substrate. The presence of M-methylamides in NRPs is believed to protect the peptide from proteolysis. Interestingly, N-MT domains are incorporated into the A domains of C-A-MT-PCP modules, between two of the core motifs (A8 and A9). MT domains contain three sequence motifs important for catalysis. ° 0-Methyl transferase domains are also found in NRPSs and likewise use the SAM cofactor. For instance, cryptophycin and anabaenopeptilide synthetases contain 0-MT domains for the methylation of tyrosine side chains. These 0-MT domains lack one of the three core motifs described for N-MT domains. ... 

Sequence Specificity Optical studies on the binding of intercalative drugs to dinucleoside monophosphates demonstrated that actinomycin exhibits a purine(3 -5 )pyrimidine specificity (57-60) while ethidium exhibits a pyrimidine(3 -5 )purine (61-63)... 

Pyrolysis-gas chromatography was used to determine the degradation of peptides to diketopiperazines (among them Pip-Sar and cis- and trans-Wal-Pip, like 48) for sequence determination in actinomycins [71JCS(CC)39],... 

For example, in cultures of Bacillus subtilis, protein synthesis had ceased entirely after one fourth of a doubling time when RNA synthesis was specifically and completely inhibited by actionomycin D33. Since the initiation of a new round of DNA biosynthesis requires the ad hoc synthesis of initiator protein(s), DNA biosynthesis comes to a standstill after a lag as the result of the failure of initiator protein synthesis. This interesting sequence of events was first described by Kirk34 for the action of actinomycin D in Staphylococcus aureus. 

Eighteen different actinomycins have been described, differing in the amino acid sequence in the peptide chains and the total synthesis of some have been reported.88 Much of our knowledge of this class is due to the work of Brockmann, who has published comprehensive reviews.47,89 Recently Weinstein et al.90 reported the synthesis of simple actinomycin analogs, which might yield effective antibacterial compounds or antitumor agents less toxic than the natural actinomycins. 

Two types of single-stranded, intramolecular triple helices of the type shown in Figure 12(d) are possible by double-hairpin formation [37]. This was demonstrated for the R RY motif by the design of suitable 18-mer sequences containing two G tracts to fold in an antiparallel fashion. The salient features of this intramolecular triplex motif are that (1) Mg2+ is not essential, (2) compared to the intermolecular version, a shorter triplex stem can be formed in a much lower DNA concentration, and (3) acidic conditions are not required as it is the R-RY motif. The evidence for triplex existence came from the observation that duplex-hairpin intercalator binders such as actinomycin D, chromomycin Aj and ethidium bromide bind poorly. 

For this review another classification is used. As the majority of peptide antibiotics are of unknown structure and cannot therefore be classified chemically, they are grouped according to the organism which produces them. At the beginning of every section, the peptide antibiotics are listed in a table which gives the known characteristics and the literature. Only the clinically useful members are described in detail. Furthermore, only those peptide antibiotics which consist mainly of amino acids in peptide linkages are fully considered. The important actinomycins , penicillins and cephalosporins, which have been reviewed several times in recent years, are not included here. Amino acids and amino acid residue sequences are denoted in accordance with the suggestions of the committee on nomenclature which reported at the Fifth European Peptide Symposium . When necessaiy the direction of the —CO NH— bond is indicated by an arrow (— )., ... 

Many other examples were then discovered. The principal androgenic steroid, dihydrotestosterone, was found to be quite specifically bound by an acidic protein in androgen-dependent tissues such as the prostate gland (Anderson and Liao, 1968). Corticosteroids, too, are bound by specific protein in the cytoplasm of liver and some other cells, and the complex enters the nucleus where it is bound by DNA. This leads to the appearance of a specific mRNA (its formation suppressable by actinomycin D) which, in turn, produces enzymes characteristic of the corticosteroid (Sekeris, 1971). A similar sequence governs the diuretic effect of aldosterone (Edelman, Bogoroch and Porter, 1963). 

Alkaline hydrolysates of mRNA from vaccinia virus (Kates, 1970) and mouse sarcoma 180 cells (Mendecki et al, 1972) show that the poly (A) tracts, 100-200 nucleotides in length, are located at the 3 -ter-minal end of the RNA molecules. Analysis of the reaction products of highly purified exoribonuclease specific for 3 -OH termini also indicates that most (and possibly all) of the poly (A) sequences are at the 3 -terminal end of the mRNA s (Molloy et al, 1972). This is supported by the fact that the time course of poly( A) labeling in polysomes indicates that this sequence is assembled after the rest of the RNA molecule has been completed (Mendecki et al, 1972). Poly (A) synthesis is sensitive to actinomycin D but to an extent less than that of the rest of the RNA molecule which further argues that the poly (A) segment is added after transcription is completed (Darnell et al, 1971b Mendecki et al, 1972). 

Nonribosomal RNA appears in the cytoplasm in the form of ribonu-cleoprotein particles called informosomes (Spirin et ah, 1964 Spirin, 1969). Such ribonucleoprotein particles, which are mostly not bound to ribosomal subunits, have been found in a variety of tissues. They are in part incorporated into polyribosomes (see Lebleu et ah, 1971 Knochel and Tiedemann, 1972 Bonanou-Tzedaki et ah, 1972). The appearance of messenger-ribonucleoprotein complexes in the polysomes is a slow process as compared to the synthesis of nuclear RNA. In chick embryos newly synthesized messenger (nonribosomal) RNA is detectable in polyribosomes after about 30 minutes. The time sequence of incorporation of mRNA into polyribosomes does not change when the RNA synthesis is completely blocked after 15 minutes by actinomycin D (Knochel, 1972b). In HeLa cells mRNA appears in the polysomes after a 15-20 minutes lag phase (Penman et ah, 1968). 

Infection with picornaviruses results in a strong (though selective) inhibition of the cellular ENA synthesis (5, 4> 85> 84), and the appearance of a new, virus-induced ENA synthesizing activity. The latter was soon shown to be insensitive to the action of Actinomycin D (5), an antibiotic which prevent DNA-dependent ENA synthesis by intercalating in the dG-dC sequences of the template MA (6, 7) These observations suggested the possibility of treating infected cultures with the antibiotic in order to suppress host-cell activity and measxiring the incorporation of radioactive precursors into viral ENA. Since the precursors added to the medium do not equilibrate instantaneously with the intracellular pool of nucleotides, a correction for this fact must be introduced, at least for the very early times. 

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