Studies on the reaction of fluorescamine with primary amines.
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In preparations of photochemical reaction centers from Rhodopseudomonas spheroides R-26, lowering the recox potential so as to reduce the primary electron acceptor prevents the photochemical transfer of an electron from bacteriochlorophyll to the acceptor. Measuring absorbance changes under these conditions, we found that a 20-ns actinic flash converts the reaction center to a new state, P-F, which then decays with a half-time that is between 1 and 10 ns at 295 degrees K. At 25 degrees K, the decay half-time is approx. 20 ns. The quantum yield of state P-F appears to be near 1.0, both at 295 and at 15 degrees K. State P-F could be an intermediate in the photochemical electron-transfer reaction which occurs when the acceptor is in the oxidized form. Following the decay of state P-F, we detected another state, P-R, with a decay half-time of 6 mus at 295 degrees K and 120 mus at 15 degrees K. The quantum yield of state P-R is approx. 0.1 at 295 degrees K, but rises to a value nearer 1.0 at 15 degrees K. The kinetics and quantum yields are consistent with the view that state P-R forms from P-F. State P-R seems likely to be a side-product, rather than an intermediate in the electron-transfer process. The decay kinetics indicate that state P-F cannot be identical with the lowest excited singlet state of the reaction center. One of the two states, P-F or P-R, probably is the lowest excited triplet state of the reaction center, but it remains unclear which one.
Lipoprotein complexes, containing (1) bacteriochlorophyll reaction centers, (2) bacteriochlorophyll light-harvesting antenna or (3) both reaction centers and antenna, have been isolated from chromatophores of non-sulphur purple bacteria Rhodospirillum rubrum by detergent treatments. The method of reconstituting the proteoliposomes containing these complexes is described. Being associtated with planas azolectin membrane, ptoteoliposomes as well as intact chromatophores were found to generate a light-dependent transmembrane electric potential difference measured by Ag/AgC1 electrodes and voltmeter. The direction of the electric field inproteoliposomes can be regulated by the addition of antenna complexes to the reconstitution mixture. The reaction center complex proteoliposomes generate an electric field of a direction opposite to that in chromatophores, whereas proteoliposomes containing reaction center complexes and a sufficient amount of antenna complexes produce a potential difference as in chromatophores. ATP and inorganic pyrophosphate, besides light, were shown to be usable as energy sources for electric generation in chromatophores associated with planar membrane.
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A new approach has been developed for the study of model and natural biomembranes. This involves the cross-linking of diacetylene groups after ultraviolet irradiation. For the study of model biomembranes, pure phospholipids (phosphatidylcholines) have been synthesized containing diacetylene groups in each acyl chain. The physical properties of these lipids have been examined and the conditions under which they polymerise have been determined. Polymerisation occurs when the lipid is in a crystalline phase, either compressed in KBr, dispersed in water (liposomes) or deposited on a suitable support (multilayers). The resultant polymer contains a conjugated backbone and is coloured. The visible spectrum of the phospholipid polymer is sensitive to its environment. Preliminary experiments show that similar polymerisation can be induced in Acholeplasma laidlawii cells grown on diacetylenic fatty acid.
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A specific exo-1,4-glucosidase (1,4-alpha-D-glucan glucohydrooase, EC 3.2.1.3) from Aspergillus niger has been partially purified and subsequently characterized by biochemical, physico-chemical and optical methods. Molecular sieve chromatography yields an enzyme with maximal activity at pH 4.2-4.5 close to its isoelectric point. Reduction and carboxymethylation leads to complete loss of activity and O-acetylation of 3 of the 13 tyrosine residues results in loss of 20 % of the activity. Sodium dodecylsulfate-polyacrylamide gel electrophoresis indicates that the native enzyme consists of two major components of molecular weights 63 000 and 57 500, respectively. Small amounts of dissociated material of molecular weight 28 000 and 16 000 as well as aggregates of the order of 100 000 are also present to the extent of 2-5% of the total potein. Following reduction and carboxymethylation under forcing conditions, the bands around 60 000 diminish and the 28 000-30 000, 16 000 and aggregate bands are dominant...
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