Changes in Ribonucleic Acid Turnover During Aerobic and Anaerobic Growth in Rhodopseudomonas spheroides

Differences in the rate and extent of degradation of ribonucleic acid (RNA) labeled by a 30-sec pulse in aerobically or anaerobically grown Rhodopseudomonas spheroides have been studied by using rifampin to block RNA synthesis. In anaerobic cultures, unstable RNA is degraded with a half-life of 1.25 to 2.0 min, and about 40% of the pulse-labeled RNA is stable. In aerobic cultures, the half-life of unstable RNA is increased to 2.5 to 4.0 min, and 50% of the RNA is stable. When aerobic cultures are transferred to anaerobic conditions, there is a rapid drop in half-life and in the proportion of stable RNA. When anaerobic cultures are made aerobic, the reverse changes occur after a lag of about 30 min. Addition of puromycin to either aerobic or anaerobic cultures caused the pulse-labeled RNA to be degraded at the same rate and to the same extent as the RNA in an anaerobic control culture. In contrast, addition of chloramphenicol enhanced the difference in RNA half-life and increased the proportion of stable RNA by about 10% in each case. It is concluded that there is a difference in the stability of an RNA component under aerobic and anaerobic conditions.     

Bacteriophages of Rhodopseudomonas spheroides: Isolation and Characterization of a Rhodopseudomonas spheroides Bacteriophage

A DNA-containing bacteriophage, designated RS1, infecting Rhodopseudomonas spheroides 2.4.1, has been isolated from sewage. The buoyant density of RS1 in CsCl equilibrium centrifugation is 1.50 g/cm(3), and the buoyant density of RS1 DNA is 1.706. The phage possesses a polyhedral head, approximately 65 nm in diameter, and a tail 60 nm long. When grown on aerobic cells, RS1 has a latent period of 120 min and an average burst size of 20. When grown on anaerobic cells, RS1 has a latent period of 150 min, and a burst size similar to that observed during aerobic infection. The adsorption rate constant of RS1 to aerobic cells is 1.2 x 10(-9) ml/min, and 0.58 x 10(-9) ml/min to anaerobic cells. Adsorption of RS1 to R. spheroides requires the presence of divalent cations.     

Ferrochelatase of Rhodopseudomonas spheroides

Extracts of Rhodopseudomonas spheroides contain two ferrochelatases: one is soluble and forms metalloporphyrins from deuteroporphyrin and haematoporphyrin; the other is particulate and forms metalloporphyrins from protoporphyrin, mesoporphyrin, deuteroporphyrin and haematoporphyrin. Neither enzyme incorporates Mg²⁺ into porphyrins or Fe²⁺ into porphyrin cytochrome c. By using the particulate enzyme, plots of 1/v versus 1/s when one substrate was varied and the other kept constant showed that neither substrate affected the K_m of the other. The suggested sequential mechanism for the reaction is supported by derivative plots of slopes and intercepts. The K_m for deuteroporphyrin was 21.3μm and that for Co²⁺ was 6.13μm. The enzyme incorporated Co²⁺, Fe²⁺, Zn²⁺, Ni²⁺ and Mn²⁺; Cd²⁺ was not incorporated and was an inhibitor, competitive with respect to Co²⁺, non-competitive with respect to deuteroporphyrin. The K_i for Cd²⁺ was 0.73μm. Ferrochelatase was inhibited by protohaem, non-competitively with respect to Co²⁺ or with respect to deuteroporphyrin. Inhibition by magnesium protoporphyrin was non-competitive with respect to deuteroporphyrin, uncompetitive with respect to Co²⁺. The inhibitory concentrations of the metalloporphyrins are lower than those required for the inhibition of δ-aminolaevulate synthetase by protohaem. Fe²⁺ is not incorporated aerobically into porphyrins unless an electron donor, succinate or NADH, is supplied; the low aerobic rate of metalloporphyrin synthesis obtained is insensitive to rotenone and antimycin. The rate of Fe³⁺ incorporation increases as anaerobic conditions are achieved.     

Phytol and Bacteriochlorophyll Synthesis in Rhodopseudomonas spheroides

Phytol has been separated and identified in the unsaponifiable lipid fraction from wild type Rhodopseudomonas spheroides, but it was not detected in mutant strains blocked at various stages of bacteriochlorophyll synthesis. Incorporation of ¹⁴C-acetate into phytol paralleled bacteriochlorophyll synthesis in suspensions of the wild type incubated anaerobically in the light. The addition of chloramphenicol inhibited both processes. It is concluded that phytol formation is tightly coupled to the synthesis of the pyrrole component of bacteriochlorophyll.     

Electron microscopy of chromatophores of Rhodopseudomonas spheroides

Gibson, K. D. (St. Mary's Hospital Medical School, London, England). Electron microscopy of Rhodopseudomonas spheroides. J. Bacteriol. 90:1059-1072. 1965.-Fixed and stained chromatophores and whole cells of anaerobically grown Rhodopseudomonas spheroides were examined in thin sections in the electron microscope. Both purified chromatophores and intracellular membrane-bound vesicles had exactly the same appearance, namely that of spheres or ellipsoids with a thin electron-dense shell surrounding an electron-lucent interior. The distribution of diameters in the two types of structure was also found to be the same, and was compatible with a normal distribution, with a mean of 570 A and a standard deviation 40 A. Negatively stained chromatophores appeared like discs or collapsed spheres. The presence of invaginations of the cytoplasmic membrane in this species was confirmed, and a new structure resembling a twin chromatophore was observed. The bearing of these results on theories of the origin of chromatophores is discussed, and it is concluded that they offer some support for each one of the three main theories about the origin of particulate organelles.     

Ribonucleic Acid from Aerobically and Anaerobically Grown Rhodopseudomonas spheroides: Comparison by Hybridization to Chromosomal and Satellite Deoxyribonucleic Acid

Ribonucleic acid (RNA) species from aerobically and anaerobically grown Rhodopseudomonas spheroides were compared via hybridization to deoxyribonucleic acid (DNA). Both long-labeled and stable RNA bound to chromosomal DNA to the same extent, regardless of derivation. About 4% of the chromosomal DNA hybridized with total cell RNA and about 0.08% with stable RNA. About 4% of the mixed satellite DNA could be hybridized to total cell RNA from aerobic or anaerobic cultures, whereas essentially no stable RNA formed a hybrid with this DNA. Hybridization competition experiments with aerobic and anaerobic pulse-labeled RNA and chromosomal or satellite DNA demonstrated that no qualitative differences existed between the RNA species. It is concluded that identical species of RNA in the same relative amounts are synthesized by R. spheroides during aerobic or anaerobic growth on the same medium.     

Mutant of Rhodopseudomonas spheroides Unable to Grow Aerobically

We isolated a mutant of Rhodopseudomonas spheroides that grows normally under photosynthetic conditions but is unable to grow exponentially under aerobic conditions. Photosynthetically grown cultures of the mutant increase in mass and cell number for about 10 hr after transfer to aerobic conditions. During this time, no heme pigments are synthesized and the Q(O(2)) declines. In the mutant, synthesis of heme pigments is obligatorily coupled to synthesis of bacteriochlorophyll.