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101.
A series of chemically-defined adenosine phosphate ligands attached to Sepharose 4B were used as active-site probes in studying the interaction of enzymes with their coenzymes and substrates and to test the suitability of these matrices for `general ligand' affinity chromatography. Nicotinamide nucleotide-dependent dehydrogenases were used as models to test this methodology. Elution from these columns by NAD+ and/or AMP gradients (in the presence or the absence of substrates and/or nicotinamide mononucleotide) was consistent with: (1) the compulsory ordered addition of substrates to lactate and malate dehydrogenase; (2) the necessity for the NMN moiety of NAD+ to bind to these enzymes before the substrate; and illustrated: (3) that the binding of these two hydrogenases to these columns compared very well with the published three-dimensional models for these enzymes and (4) that separation of mixtures of dehydrogenases depended on the choice of matrix and displacing ion and whether any additions (e.g. substrates) were made to the gradients used. These techniques were used to purify UDP-glucose dehydrogenase from a crude starting material on a phosphate-linked UDP (or ADP) matrix. The binding of this enzyme to these two columns was not consistent with either an ordered or random addition of substrates and suggested a more complex mechanism.  相似文献   
102.
The methods of Atkins & Nimmo (1973) and Fernley (1974) for fitting the integrated Michaelis-Menten equation were compared by using the same sets of simulated experimental data. The method of Fernley (1974) is to be preferred because it gives precise and unbiased estimates of the Michaelis-Menten parameters over a wide range of substrate concentrations. However, the estimates may not be symmetrically distributed, especially at low substrate concentrations.  相似文献   
103.
Arnold Medved  Ian Maxwell 《CMAJ》1974,111(3):245-250
Cyclophosphamide, given in widely spaced doses, was used in the treatment of a patient with pemphigus vulgaris and a patient with bullous pemphigoid. To our knowledge, this form of therapy has not previously been reported in these two diseases. The distinct advantages of the larger intermittent dose method of cyclophosphamide therapy over the more conventional daily dose regimen are discussed.  相似文献   
104.
To assess the possible role of filaments in subcellular motility, particular cultured cells were studied by light and electron microscopy. Motile cell margins always contained meshworks of ~50 Å diam. filaments. Organelles moved within cytoplasm occupied by a meshwork of 50–100 Å filaments and microtubules. When cells were treated with cytochalasin B, movements of cell margins stopped, but organelle movements continued; electron microscopically, while subplasmalemmal filaments had disappeared, subcortical filaments and microtubules remained. When cells were treated with hypertonic medium, organelle movements ceased but marginal movements continued; electron microscopically, although cell margins contained normal filament arrays, few subcortical filaments remained. It is concluded that while cell margins are moved by a meshwork of filaments, organelle movement is mediated by a subcortical meshwork of filaments and microtubules.  相似文献   
105.
6-MSA3 synthase has been purified 190-fold with 33% yield. The purification was found to be dependent on the presence of glycerol. The acetylenic inhibitors 3-pentynoyl- and 2-hexynoyl-NAC completely inhibit 6-MSA production at concentrations in which fatty acid synthesis, TAL production as well as NADPH oxidation are only partially affected. These results confirm earlier studies on the specificity of inhibition by acetylenic inhibitors and support a mechanism wherein the NADPH-mediated reduction step occurs on a 6-carbon rather than on an 8-carbon intermediate.  相似文献   
106.
1. The preparation of enzymically active N-citraconyl derivatives of fructose diphosphate aldolase from rabbit muscle is described. Reaction is restricted to amino groups and the derivatives are not very heterogeneous with respect to the number of substituents. 2. Linear double-reciprocal plots of enzyme velocity against substrate concentration are found up to about 15% blocking of amino groups. With more than 15% blocking, there is a marked downward curvature in the double-reciprocal plots at high substrate concentrations. 3. Over the range 0-25% blocking of amino groups the apparent V(max.) for fructose diphosphate falls to 10% that of the native enzyme, and the apparent K(m) rises from 1 to 400mum. 4. Various pieces of evidence suggest that citraconyl-aldolase is slightly distorted in structure compared with the native enzyme. However, the kinetic properties and tetrameric structure of citraconyl-aldolase can be completely recovered after denaturation in 4m-guanidine hydrochloride. 5. After removal of the citraconyl groups in acid conditions the kinetic and molecular properties of native enzyme are restored. 6. Hybrid forms of aldolase can be constructed containing native and citraconylated subunits and the suitability of these derivatives for the study of subunit interactions in the enzyme is discussed. 7. The kinetic properties of hybridized aldolase containing native and citraconylated subunits are not exactly those predicted from the kinetic properties of the two parental forms. This result is interpreted in terms of conformational changes induced in the native and modified subunits when both are present in a hybrid molecule, evidently as a result of interactions in the tetramer.  相似文献   
107.
The vascular bundle sheath cells of sugar cane contain starch-storing chloroplasts lacking grana, whereas the adjacent mesophyll cells contain chloroplasts which store very little starch and possess abundant grana. This study was undertaken to determine the ontogeny of these dimorphic chloroplasts. Proplastids in the two cell types in the meristematic region of light-grown leaves cannot be distinguished morphologically. Bundle sheath cell chloroplasts in tissue with 50% of its future chlorophyll possess grana consisting of 2-8 thylakoids/granum. Mesophyll cell chloroplasts of the same age have better developed grana and large, well structured prolamellar bodies. A few grana are still present in bundle sheath cell chloroplasts when the leaf tissue has 75% of its eventual chlorophyll, and prolamellar bodies are also found in mesophyll cell chloroplasts at this stage. The two cell layers in mature dark-grown leaves contain morphologically distinct etio-plasts. The response of these two plastids to light treatment also differs. Plastids in tissue treated with light for short periods exhibit protrusions resembling mitochondria. Plastids in bundle sheath cells of dark-grown leaves do not go through a grana-forming stage. It is concluded that the structure of the specialized chloroplasts in bundle sheath cells of sugar cane is a result of reduction, and that the development of chloroplast dimorphism is related in some way to leaf cell differentiation.  相似文献   
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