Biochimica et Biophysica Acta (BBA) - Specialized Section on Biophysical Subjects最新文献

Addition of H₂O₂ to a bacterial luciferase preparation resulted in the formation of a compound with a fluoresence maximum at about 470 mμ. This compond was probably the same as the one that is formed as a result of ultraviolet irradiation of luciferase preparations after addition of FMNH₂. It is assumed to be the luminescent molecule in bacterial luminescence.

The effect of a number of substances (K₃Fe(CN)₆, K₄Fe(CN)₆, ascorbic acid, peroxidase (donor: H₂O₂ oxidoreductase, EC 1.11.1.7), tryptophan and p-chloromecuribenzoate) on both the light reaction and fluorescence increase reaction was investigated. Substances inhibiting the fluorescence increase reaction (K₃Fe(CN)₆, K₄Fe(CN)₆, ascorbic acid) likewise could inhibit the light reaction. Both effects are, at least partly, ascribed to an effect on the fluorescent group of luciferase. Light reaction inhibitors affecting other groups at the enzyme surface (p-chloromercuribenzoate) did not influence the fluorescence increase effect. Activators of the light reaction (cystine, K₄Fe(CN)₆) are suggested to act by their protective action on sulfhydryl groups or on dissociable groups, presumed to be attached to these sulfhydryl groups.

A discussion of the results leads to the conclusion that the light reaction consists of at least two consecutive reactions, involving two different groups of the luciferase molecule.

Quantitative determinations were made of the number of spins responsible for the sharp, fast decaying EPR signal induced by light in photosynthetic material. The species resposible for the observed EPR signal is present at the level of approx. 1 per 100 molecules of chlorophyll. Values ranging between 1 and 4 were obtained for the ratio between free radicals and molecules of photoconverter P700.

In view of the difficulties and errors inherent in the quantitation procedures involved, this finding does not exclude the possibility that the oxidized of P700 is the species responsible for the observed EPR signal, as the correlation of several properties of the radical and P700 had previously suggested. The presence of radical species other than or in addition to P700⁺ is, however, also compatible with the results.

• 1.

  1. The spectral dependence of dichroism of chloroplasts in vivo was measured in order to determine if chloroplast bifluorescence is due to an oriented absorbing pigment system.

• 2.

  2. Dichroic ratios of 2.5–4.0 were obtained in the spectral region of 705 mμ where dichroism is maximum.

• 3.

  3. The direction or sense of the dichroism in chloroplasts indicates that the absorbing oscillator lies in or near the plane of the chloroplast lamellae. This is an accord with out previous observations of bifluorescence.

• 4.

  4. No dichroism of chloroplasts in vivo could be detected in the ultraviolet spectral region from 250 to 400 mμ nor in the visual region from 400 mμ to 570 mμ.

• 5.

  5. A consideration of the pigment orientation in terms of energy transfer and in terms of macromolecular complexes is offered in a discussion involving the comparison of chromophore orientation in analogous paracrystalline systems both in vivo and in vitro.

• 1.

  1. Spectral measurements of the bifluorescence of chloroplasts in vivo indicate a high degree of orientation of a far-end pigment having a maximum emission near 716 mμ.

• 2.

  2. This oscillator of emission lies in or near the plane of the chloroplast lamellae and its existence, in some way, depends upon the structural integrity of the chloroplast at the molecular level.

• 3.

  3. The chromophore involved is strategically located to accept excitation energy from ordinary chlorophyll or accessory pigments, and the form of its molecular organization may be common with that of other photobiological systems.

The behaviour and physico-chemical characteristics of corticotropin, or adrenocorticotropic hormome (ACTH), in aqeous solution as a function of ionic strength and temperature have been studied. It is shown that sharp changes of physico-chemical properties of the solution take place when both temperature or ionic strength reach critical values. Such changes are discussed in terms of structural variations of the ACTH molecule.

Electron transfer as opposed to hydrogen transfer was demonstrated to be involved in the oxidation-reduction of the flavoprotein enzyme system. A bioelectrochemical investigation of glucose oxidase (EC 1.1.3.4), d-amino acid oxidase (EC 1.4.3.3), and yeast alcohol dehydrogenase (EC 1.1.1.1) systems was conducted in an attempt to utilize the electron-transferring process as a potential anodic reaction in a biochemical fuel cell. Utilizing a bio-fuel cell constructed of plexiglass, platinum-foil electrodes, and an ion-exchange membrane for conduction between the anolyte and catholyte, the flavoprotein enzymes, both glucose oxidase and d-amino acid oxidase systems in conjunction with an O₂ cathode, generated 175–350 mV. In contrast, alcohol dehydrogenase (yeast), a pyridinoprotein enzyme which requires coenzyme I (NAD+), did not produce any electrical voltage. Elemental iron was found to potentiate the flavoprotein enzyme reaction yielding voltages ranging from 625 to 750 mV. The potentiating effect was probably due to a faster turnover rate of FADH to FAD+ coupled with the additional net oxidation potential of iron.

• 1.

  1. Mutants of Rhodopseudomonas spheroides have been isolated that cannot grow photosynthetically but can grow aerobically (in the dark) at the same rate as the parent strain. The major pigments of the photosynthetically dificient mutant PM-8 and its parent strain Ga are bacteriochlorophyll, neurosperene, and hydroxyneurospene. Under conditions of limited aeration both strains contain very similar amounts of these pigments. In chromatophore fractions from these two strains the bacteriochlorophyll shows the same degree of susceptibility to structural and photo-oxidative alterations in the presence of detergents.

• 2.

  2. A blue-green mutant strain, PM-8:bg58, derived from strain PM-8, suffers photooxidative injury sensitized by bacteriochlorophyll in the same way as do photosynthetically competent blue-green (colored carotenoid-less) strains such as uv-33.

• 3.

  3. Strain PM-8 exhibits none of the reversible light-induced absorbancy changes detectable in strain Ga. These changes, some of which have been implicated in the primary photochemistry of photosynthesis, reflect oxidation of cytochromes, oxidation of P870 (a specialized, minor bacteriochlorophyll component), reduction of ubiquinone, and a shift in the absorption bands of carotenoids. A chemically induced difference spectrum characteristics of the oxidation of P870 is also lacking in strain PM-8.

• 4.

  4. Strain PM-8 does not show a light-induced electron-spin-resonance signal, whereas photosynthetically competent strains of purple bacteria do.

• 5.

  5. The yield of bacteriochlorophyll fluorescence in strain PM-8 is about two times that in strai Ga. A transient increase in flourescence intensity correlated with bleaching of P870 was observed in strain Ga but not in strain PM-8.

• 6.

  6. It is concluded that in strain PM-8 the photochemical reaction centers that initiate the chemistry of photosynthesis are absent or inoperative.

Iron-containing proteins present in Euglena have been examined. The properties of pure samples of two of these—cytochrome c (552, Euglena) and cytochrome c-type (558, Euglena)-have been determined in some detail. These include spectrochemical characteristics, reactivity, stability, amino acid composition, isoelectric point, midpoint potential and nature of haem prosthetic group, in addition to molecular weight and haem content. A third haem protein, particle bound, and associated with the chloroplast fraction, is characterized as a cytochrome b₆. Evidence for the existence of an a-type cytochrome and ferrodoxin is also presented.