Hiroshi Yamaki

The combined effects of bleomycin and sulfhydryl compounds on the thermal denaturation of DNA.

Abstract The reaction of bleomycin with DNA in vitro was shown to occur by a decrease in the melting temperature ( T m of DNA. This decrease of T m was observed when DNA was incubated with the antibiotic in the presence of a sulfhydryl compound such as 2-mercaptoethanol or dithiothreitol. It was also observed when DNA was incubated with 2-mercaptoethanol, dialyzed and thereafter bleomycin A 2 was added. It was not observed when DNA was incubated with bleomiycn A 2 alone and with the addition of 2-mercaptoethanol just before the determination of T m . The results indicate that, although 2-mercaptoethanol has no effect on T m , DNA must react with 2-mercaptoethanol since the T m of DNA after the reaction is decreased, as shown by the reaction with bleomycin A 2 . The effect of bleomycin A 2 on DNA in the presence of the sulfhydryl compound was diminished by the addition of Cu 2+ , Zn 2+ or Co 2+ . Bleomycin mixture or bleomycin A 2 in the copper-containing form does not significantly affect T m . Bleomycins and phleomycins are closely related peptide antibiotics, but phleomycin increased the T m of DNA irrespective of the presence of the sulfhydryl compound.

The mechanism of antifungal action of (S)-2-amino-4-oxo-5-hydroxypentanoic acid, RI-331: the inhibition of homoserine dehydrogenase in Saccharomyces cerevisiae.

We have explored the mechanism by which an antifungal antibiotic, (S)-2-amino-4-oxo-5-hydroxypentanoic acid, RI-331, preferentially inhibits protein biosynthesis in Saccharomyces cerevisiae, by inhibiting the biosynthesis of the aspartate family of amino acids, methionine, isoleucine and threonine. This inhibition was effected by inhibiting the biosynthesis of their common intermediate precursor homoserine. The target enzyme of RI-331 was homoserine dehydrogenase (EC.1.1.1.3) which is involved in converting aspartate semialdehyde to homoserine in the pathway from aspartate to homoserine. The enzyme is lacking in animals. So the antibiotic is selectively toxic to prototrophic fungi.

Inhibition of initiation of DNA synthesis by aminoglycoside antibiotics

In a dnaCts mutant of E. coli, the reinitiation of DNA synthesis, which occurred by the shift of the culture from a restrictive temperature to a permissive temperature, was markedly prevented by habakacin, dibekacin, kanamycin, and gentamicin. On the contrary, chloramphenicol did not inhibit the reinitiation synthesis for 30 min. In a parallel experiment, leucine uptake into protein was profoundly blocked by chloramphenicol, but only slightly by habekacin. Habekacin did not significantly affect DNA elongation of the cells at a restrictive temperature. We propose that inhibition of initiation of replication by aminoglycoside antibiotics is related to their lethality.

Selective inhibition of initiation of globin synthesis by phenomycin

Summary It has been demonstrated in a rabbit reticulocyte system that phenomycin selectively blocks the initiation of globin synthesis. The N-terminal incorporation of valine or methionine is more markedly inhibited by the antibiotic than the total incorporation. The 80S initiation complex is accumulated in the presence of phenomycin. The initial dipeptide formation (Met-Val) is significantly affected by the antibiotic.

A peplomycin-supersensitive cell line lacking activation of poly(adenosine diphosphate ribose) synthetase by peplomycin

In peplomycin-supersensitive Chinese hamster lung cells, the increase in poly(ADP-ribose) synthesizing activity following peplomycin treatment was significantly reduced as compared with the parental lung cells, suggesting that peplomycin-supersensitive lung cells may have some deficiency in DNA repair. On the contrary, peplomycin-supersensitive ovary cells, which undergo increased DNA damage induced by peplomycin, showed normally increased poly(ADP-ribose) polymerizing activity compared with the parental ovary cells. Relationship between poly(ADP-ribose) polymerase and peplomycin sensitivity was discussed.

Functional interaction of free polyribosomes with the membrane of the endoplasmic reticulum in a cell-free protein-synthesizing system from plasmacytoma X5563

Abstract In cell-free protein synthesis by the murine plasmacytoma X5563, which had become a nonproducing mutant, mixed systems with free polyribosomes and mirosomes incorporated 14 C-amino-acid into protein 3–8 times greater than the sum of the incorporations in the individual system irrespective of S-100 concentrations. This enhancement was inhibited by lecithinase A and was markedly reduced at high KCl concentrations. Smooth endoplasmic membranes had more stimulatory activity than rough endoplasmic membranes. The results indicate that the membrane of the endoplasmic reticulum and free polyribosomes interact in the cell-free protein-synthesizing system, resulting in the enhancement of protein synthesis.