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

• 1.

  1. With chromatophores from Rhodospirillum rubrum, phosphorylations coupled with photosynthetic electron-transport systems have been investigated.

• 2.

  2. Under illumination with white light at a fixed high intensity, the addition of a strictly limited concentration of phenazine methosulfate was required to induce photophosphorylation at a maximal rate. The maximal rate of photophosphorylation was always much faster when induced by phenazine methosulfate than when induced by the other reagents tested such as ascorbate or succinate. The ratio of maximal rate of the phenazine methosulfate-induced to ascorbate-induced photophosphorylation varied from batch to batch of chromatophore preparations with experimental values from 2 to 5.

• 3.

  3. The phenazine methosulfate-induced, and ascorbate-induced photophosphorylations were inactivated in a similar manner by illumination of chromatophores with ultraviolet irradiation and by additions of the following inhibitors: Na₂SO₄, quinacrine hydrochloride, 2,4-dinitrophenol, dicumarol EDTA and phenylmercuric acetate. Chromatophore suspensions, if resonicated or heated, lost the activity of ascorbate-induced photophosphorylation somewhat more rapidly than that of phenazine methosulfate-induced photophosphorylation. In the presence of an appropriate concentration of o-phenanthroline, the ascorbate-induced and succinate-induced photophosphorylations were inactivated completely, but the phenazine methosulfate-induced photophosphorylation was not affected. With the chromatophores treated with an appropriate concentration of Triton X-100, the ascorbate-induced and succinate-induced photophosphorylations were inactivated completely, whereas the phenazine methosulfate-induced photophosphorylation was inactivated only partially.

• 4.

  4. The photooxidations of cytochrome c₂, and cytochrome c were stimulated in rate by the addition of ADP. The stimulation by ADP was not significantly influenced in the presence or absence of P_i. The photooxidation was stimulated by 2,4-dinitrophenol in the absence but not in the presence of ADP. The concentraton of 2,4-dinitrophenol required for the stimulation was nearly identical with that required for the inhibition of photophosphorylation.

• 5.

  5. ATP was synthesized from ADP and P_i, coupled with the photooxidations of cytochrome c₂, and cytochrome c. The values of the P/O ratio obtained were less than one.

• 6.

  6. Based on these findings, the site for phosphorylation coupled with the photosynthetic electron-transport systems are suggested.

In order to differentiate between the various proposed models for the structure bacterial flagella, low-angle X-ray diffraction studies have been carried out on wet specimens of Proteus vulgaris. The low-angle equatorial diffraction pattern has been obtained and compared with the theoretically calculated patterns for a rod model and the double and triple helical structures according to Labaw and Mosley. The comparison shows that a model comprised of helical filaments is the most likely structure. Fourier-Bessel transformation of the experimental scattering data has been performed and the radial density-distribution curve gives low values in the middle of the structure, suggesting the flagellum has a hollow center, thus favouring the triple helical form.

• 1.

  1. Employing hamster everted intestinal sacs, the transport of l-phenylalanine and l-proline against a concentration gradient had the following indentical characteristics: spatial distribution of transport, inhibition by absence of Na+ from the medium, inhibition by NaCN but not by propionitrile, lack of effect of added ascorbic acid, and the same temperature coefficient (2.3 for a 10° change), In addition, there was mutual inhibition of transport as measured by the equillibrium quantity. These parameters, hence, were unable to provide a distinction between the mechanism of transport of the amino acid and the amino acid (although genetic data suggest that two distinct, but overlapping, transport systems are involved).

• 2.

  2. A series of noncyclic imino acids of the form

  was also studied for transport against a concentration gradient by the hamster everted small intestine. When X = H, transport occurred with Y = CH₃, but not when Y = CH₂CH₃ or larger groupings. Hence, N-methylglycine likely represents the most “sterically bulky” of these compounds to be transported.

• 3.

  3. Cyclic imino acids of the form

  were transported against a concentration gradient when n = 2 (azetidine-2-carboxylic acid), n = 3 (proline) and n = 4 (pipecolic acid). For n = 1 (aziridine-2-carboxylic acid, lithium salt), the compound did not appear to be transported. Data are presented that cyclic and noncyclic imino acids show cross-inhibition of transport.

The object of this investigation was to study the modification of histidine residues by N-bromosuccinimide leading to the appearance of visible fluorescence.

• 1.

  1. Histidine and tryptophan yield visible fluorescence when treated with N-bromosuccinimide in phosphate buffer at neutral pH. Under the conditions required for the formation of the fluorescent products, tryptophan and tyrosine lose their native ultraviolet fluorescence.

• 2.

  2. Details are presented in peptides yield mmore fluorescence than histidine itself. The amount of fluorescence formed differs from one peptide to another depending on the amino acids adjacent to the histidine residue.

• 3.

  4. The reaction leading to the formation of fluorescent products from histidine residues can experimentally be divided into two distinct steps: (a) Rapid attack of the histidine residue by N-bromosuccinimide to yield a non-fluorescent intermediate. (b) Conversion of the intermediate into a fluorescent compound in a slow reaction cataylsed by phosphate (or pyrophosphate, or hydroxyl ions). Optimal reaction conditions for both steps were determined.

• 4.

  5. The formation of fluorescent products from histidine residues when treated with N-bromosuccinimide may be of use in following the modification of histidine residues in peptides and in proteins. It may also be a means of introducing visible fluorescence into proteins for structural studies.

• 5.

  6. The possible relevance of this work to the problem of oxidative phosphorylation is briefly discussed.

The hemoglobin of red cell hemolysates of chicken embryos and chickens after hatching have been reinvestigated using starch-gel electrophoresis and O₂ equilibrium measurements. Thee hemoglobin components were found, namely the major and minor hemoglobin fractions also present in the adult chicken red cell hemolysates and a slow-moving fraction, which disappeared gradually from the blood after hatching. The electrophoretic mobility of the minor hemoglobin fraction in embryonic red cell hemolysates was slower than that of the fractions observed in 1-day-old chick and in adult chicken hemolysates, and was the same as that produced by preincubation of these hemolysates with 0.2 M phospahte buffers. It was also observed that the erythrocytic inorganic phosphate level was several times higher in the newborn chicken than in the adult fowl; the decrease in phosphate level after hatching coincides with the decrease in O₂ affinity of total blood samples as observed by others. The equilibrium of the total red cell hemolysate of a 1-day-old chicken with O₂ showed a higher O₂ affinity at low phosphate molarity, which decreased with increased phosphate concentration in a similar ways as observed for adult red cell hemolysates being pre-exposed to phosphate. The possible role of the phospahte content of the red cell in the regulation of the O₂ affinity of the chicken hemoglobin during development has been discussed.

• 1.

  1. Fructose complexes of Cu²⁺ and Fe³⁺ have been studied by ESR, NMR and magnetic susceptibility measurements. In the case of Fe³⁺, the experimental parameters have been studied in pH-titration experiments. Ultracentrifugation studies have also been performed in order to study the size of colored species in the Fe³⁺ solutions.

• 2.

  2. With Cu²⁺, the ESR spectra at room temperature as well as at 77 °K indicate the presence of a well-defined complex with approximately axial symmetry around the metal ion. The small line width at 77 °K allows resolution of hyperfine structure in the perpendicular direction also. The room temperature ESR spectrum and the NMR line width are consistent with the formation of a complex consisting of 1 Cu²⁺ ion and 2 fructose molecules.

• 3.

  3. With Fe³⁺, the ESR signal intensity, NMR line width and magnetic susceptibility go through a minimum at pH7 as increasing amounts of NaOH are added to an acid solution of the metal ion in the presence of excess fructose. The results are consistent with the formation of a dimer (consisting of 2 fructose molecules and 2 Fe³⁺ ions joined by oxygen bridges), which breaks up at higher pH values. However, the ultracentrifugation experiments indicate the presence of larger particles, be aggregates of the dimer.

The equilibrium of dilute and concentrated adult chicken red blood cell hemolysates with O₂ has been studied using potassium phosphate buffers and NaCl solutions of different molarities. The results indicate an increase in O₂ affinity with increase in ionic strength. The same phenomenon was observed for the isolated minor hemoglobin component; the isolated major hemoglobin component showed a rather high O₂ affinity at low ionic strength, which decreased with increase in molarity. The minor hemoglobin fraction exhibited an ability to form rather stable complexes with phosphate ions; such complexes possessed a high O₂ affinity at low ionic strength. The data also indicated that the two hemoglobin components in a hemolysate interacted with each other. This extra-molecular interaction interfered with the physiologic properties of the hemolysate. No physico-chemical interaction could be detected, as was demonstrated by the determination of the molecular weight and by viscosity measurements.