Glycerolipid labelling kinetics in isolated intact chloroplasts.

Glycerolipid synthesis was studied in intact chloroplasts isolated from three different plant species. The sequential acylation of sn-glycerol 3-phosphate and lysophosphatidate (1-acyl-sn-glycerol 3-phosphate) was confirmed by monitoring the incorporation of oleate synthesized in situ into lysophosphatidate, phosphatidate and diacylglycerol. Lysophosphatidate was not only readily detected in these experiments, but was also present in the chloroplasts at the beginning of the time courses. The rate of glycerolipid synthesis depended primarily on sn-glycerol 3-phosphate supply, and given adequate sn-glycerol 3-phosphate, the proportion of newly synthesized fatty acids diverted into glycerolipids appeared to be determined by differing acyltransferase activities in the chloroplasts isolated from different plant species.     

Protein synthesis by isolated chloroplasts

Isolated chloroplasts show substantial rates of protein synthesis when illuminated. This in organello protein synthesis system has been advantageously utilised to elucidate the coding capacity of chloroplast and the regulation of chloroplast genes. The system is also being used recently to transcribe and translate homologous and heterologous templates. In this mini-review, we attempt to critically ecaluate the available literature and present the current and the prospective lines of research.     

Photoswelling and light-inactivation of isolated chloroplasts I. Change in lipid content in light-aged chloroplasts

The formation of lipid peroxide and changes in the lipid compositionof isolated chloroplasts aged in the light or dark, were investigatedin more detail. Lipid peroxide formation was observed in thethylakoid membrane as well as in the supernatant from dark-agedchloroplasts. Light was necessary for its formation in bothsystems. We confirmed that the peroxidation of lipids formedduring aging did not induce the inhibition of photochemicalactivities in chloroplasts. Aged chloroplasts underwent decompositionof their endogeneous monogalactolipid and phosphatidylcholine(lecithin) resulting in free fatty acids and lysophosphatidylcholine(lysolecithin). Decomposition of monogalactolipid occurred inboth the light- and dark-aged chloroplasts. The change of lecithinto lysolecithin was stimulated by illumination. This suggeststhat the peroxidation of lipids occurs as a result of the illuminationof free fatty acids released from monogalactolipid and lecithinin the thylakoid membranes, and that the change of lecithinto lysolecithin is related to the inactivation of photochemicalactivities and to swelling in light-aged chloroplasts. ¹ Present address: Department of Microbiology, Ishikawa ResearchLaboratory for Public Health and Environment, Minma, Kanazawa,Japan. (Received August 15, 1974; )     

Photoswelling and light-inactivation of isolated chloroplasts II. Functional and structural changes in isolated chloroplasts under the influence of lysolecithin

Lysolecithin was added to spinach chloroplasts in suspension,and its effect as a detergent on structure and electron transportactivities was examined. At low concentrations of lysolecithin,the activities of FeCN photoreduction and O₂-linked DCPIPH₂photo-oxidation were stimulated but the photoreduction of NADPwas not enhanced and decreased with an increase in concentration.Absorption bands over the whole visible region were intensifiedwithout any shifts. At high concentrations, the activities ofFeCN photoreduction and O₂-linked DCPIPH₂ photo-oxidation decreasedfrom a maximum to 10–50% of the control activity, andNADP photoreduction was completely inhibited. Absorption bandswere further intensified and the red band was slightly shifted.Results indicate that lysolecithin treatment is useful in chloroplastbiochemistry. Lysolecithin formation and the deterioration ofchloroplasts during light-aging is also discussed. (Received August 15, 1974; )     

Enhancement studies on algae and isolated chloroplasts. Part II. Enhancement of oxygen evolution in intact chloroplasts.

Intact isolated chloroplasts were shown to exhibit a characteristic three-phase pattern of development of oxygen evolution activity. The first phase, Phase I, appeared to be an equilibration phase in which the isolated chloroplasts adapted to the conditions on the electrode surface. It was characterised by a rapidly increasing rate of oxygen evolution accompanied by decreasing enhancement signals. The second phase, Phase II, was an intermediate phase in which the rate of oxygen evolution was maximal and no enhancement was observed. In the last phase, Phase III, the rate of oxygen fell again, normal enhancement was still missing, but the samples appeared to undergo slow adaptive changes closely related to the State I-State II changes previously reported for whole cell systems. The concentrations of Mg2+ within the chloroplast were shown to play an important role in the control of the development of both the oxygen evolution and enhancement signals. It was shown how these signals could be explained in terms of a model that was consistent with that developed in Part I of this investigation to account for the variability of enhancement of the alga Chlorella pyrenoidosa.     

Studies on the slow fluorescence decline in isolated chloroplasts.

Data presented here indicate that the slow fluorescence decline in osmotically disrupted chloroplasts is not associated with the well known divalent cation effect on fluorescence yield. Thus the two phenomena have markedly different magnesium concentration requirements, magnesium addition after the fluorescence decline did not stimulate the dark reversal, and the characteristics of the fluorescence induction kinetics of the two processes are not similar. At pH 7.6 the slow fluorescence decline was stimulated by several uncouplers demonstrated to greatly reduce proton pumping, and at pH 9.2 it was stimulated by all uncouplers tested. Acid-base transition was strongly inhibitory, and this inhibition was relieved by coupling factor is suggested by experiments in which phosphorylation substrates were inhibitory, and this inhibition was prevented by uncoupler. These data are explained in terms of coupling factor structural changes which in an unknown manner influence Photosystem II fluorescence emission. Fluorescence induction curves indicate that the slow quenching decreased only the variable fluorescence. The half rise time was decreased along with the sigmoidicity of the rise curve. These data can be accomodated in terms of a model recently proposed by Butler and Kitajima (Biochim. Biophys Acta (1975) 376, 116-125), involving the transfer of energy from the excited, but closed, reaction centres II to the light harvesting chlorophyll system. The slow fluorescence decline is suggested to represent a decrease of this process.