PubMed · 9664468
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E de Kant. 1998. Gene expression analysis by CD-RT-PCR.. https://doi.org/10.1385/0-89603-494-1%3A173
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A photosystem II preparation from the thermophilic cyanobacterium Synechococcus elongatus, which is especially suitable for three-dimensional crystallization in a fully active form was developed. The efficient purification method applied here yielded 10 mg of protein of a homogenous dimeric complex of about 500 kDa within 2 days. Detailed characterization of the preparation demonstrated a fully active electron transport chain from the manganese cluster to plastoquinone in the Q(B) binding site. The oxygen-evolving activity, 5000-6000 micromol of O(2)/(h.mg of chlorophyll), was the highest so far reported and is maintained even at temperatures as high as 50 degrees C. The crystals obtained by the vapor diffusion method diffracted to a resolution of 4.3 A. The space group was determined to be P2(1)2(1)2(1) with four photosystem II dimers per unit cell. Analysis of the redissolved crystals revealed that activity, supramolecular organization, and subunit composition were maintained during crystallization.
A high molecular model compound of polycyclic aromatic hydrocarbon was synthesised by coupling pyrene to PEG(5000). The pyrene-PEG was used for the study of a laccase-mediator-system. To prevent direct contact between the substrate and the enzyme the two were kept in their own compartments separated by a membrane. The low molecular mediators, 1-hydroxybenzotriazole and 2, 2'-azino-bis-(3-ethylbenzothiazoline-6-sulphonic acid), which were oxidised by laccase to the corresponding radicals or cations permeated the membrane and reacted with the pyrene-PEG model compound. Oxidation of the model compound resulted in an alpha-oxidation of the alkyl-chain leading to two main oxidation products. The same oxidation products were obtained in the reaction system without a membrane.
An LC/MS-based method is established for the differentiation and authentication of specimens and commercial samples of Panax ginseng (Oriental ginseng) and Panax quinquefolius (American ginseng). This method is based on the separation of ginsenosides present in the ginseng methanolic extracts using high-performance liquid chromatography (HPLC), followed by detection with electrospray mass spectrometry. Differentiation of ginsenosides is achieved through simultaneous detection of intact ginsenoside molecular ions and the ions of their characteristic thermal degradation products. An important parameter used for differentiating P. ginseng and P. quinquefolius is the presence of ginsenoside Rf and 24-(R)-pseudoginsenoside F11 in the RICs of Oriental and American ginsengs, respectively. It is important to stress that ginsenoside Rf and 24(R)-pseudoginsenoside F11, which possess the same molecular weight and were found to have similar retention times under most LC conditions, can be unambiguously distinguished in the present HPLC/MS method. The method developed is robust, reliable, reproducible, and highly sensitive down to the nanogram level.