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Donal S O'Leary 《Journal of applied physiology》2006,100(1):357-8; discussion 360-1
4.
1-Carboxyallenyl phosphate, the allenic homologue of phosphoenolpyruvate, has been synthesized in six steps. The key step in the synthesis is the isomerization of methyl 2-hydroxy-3-butynoate to the corresponding allenol and phosphorylation of this material. The allene is an excellent substrate for pyruvate kinase, undergoing reaction at more than half the rate of phosphoenolpyruvate. The allene is also a substrate for phosphoenolpyruvate carboxylase, being hydrolyzed by the enzyme rather than carboxylated. With both enzymes, the organic product is 2-oxo-3-butenoate, which gradually inactivates the enzymes by reaction with one or more sulfhydryl groups not at the active site. 相似文献
5.
Formation of distinct multicellular aggregates is one of the phenomena associated with activation of quiescent human mononuclear leukocytes in vitro. Aggregate formation involves active cell motility and enhances cell-cell interactions required for an optimal proliferative response of T-cells stimulated with agents like phytohemagglutinin. We have developed an assay to quantitate the rate at which motile cells form aggregates on a flat surface. This assay follows the time rate of deviation of cells in undisturbed culture away from an initial random distribution using an "aggregation index." We used this assay to establish minimal culturing conditions required to observe an aggregation response for a partially purified mononuclear leukocyte population. We also studied the ability to aggregate of various subpopulations enriched for T- and B-lymphocytes and monocytes and found evidence for a monocyte requirement for lymphocyte aggregation. In a second assay, we followed the rate of entry of esterase positive monocytes into aggregates and compared this to the rate of entry of mononuclear cells in toto. We found that monocytes are preferentially associated with non-esterase positive cells within one hour of PHA stimulation. The results support the conclusion that monocytes play a central role in directing the motility of human T-lymphocytes leading to their aggregation response in tissue culture. 相似文献
6.
C. B. Osmond J. A. M. Holtum M. H. O'Leary C. Roeske O. C. Wong R. E. Summons P. N. Avadhani 《Planta》1988,175(2):184-192
The labeling patterns in malic acid from dark 13CO2 fixation in seven species of succulent plants with Crassulacean acid metabolism were analysed by gas chromatography-mass spectrometry and 13C-nuclear magnetic resonance spectrometry. Only singly labeled malic-acid molecules were detected and on the average, after 12–14 h dark 13CO2 fixation the ratio of [4-13C] to [1-13C] label was 2:1. However the 4-C carboxyl contained from 72 to 50% of the label depending on species and temperature. The 13C enrichment of malate and fumarate was similar. These data confirm those of W. Cockburn and A. McAuley (1975, Plant Physiol. 55, 87–89) and indicate fumarase randomization is responsible for movement of label to 1-C malic acid following carboxylation of phosphoenolpyruvate. The extent of randomization may depend on time and on the balance of malic-acid fluxes between mitochondria and vacuoles. The ratio of labeling in 4-C to 1-C of malic acid which accumulated following 13CO2 fixation in the dark did not change during deacidification in the light and no doubly-labeled molecules of malic acid were detected. These results indicate that further fumarase randomization does not occur in the light, and futile cycling of decarboxylation products of [13C] malic acid (13CO2 or [1-13C]pyruvate) through phosphoenolpyruvate carboxylase does not occur, presumably because malic acid inhibits this enzyme in the light in vivo. Short-term exposure to 13CO2 in the light after deacidification leads to the synthesis of singly and multiply labeled malic acid in these species, as observed by E.W. Ritz et al. (1986, Planta 167, 284–291). In the shortest times, only singly-labeled [4-13C]malate was detected but this may be a consequence of the higher intensity and better detection statistics of this ion cluster during mass spectrometry. We conclude that both phosphoenolpyruvate carboxylase (EC 4.1.1.32) and ribulose-1,5-biphosphate carboxylase (EC 4.1.1.39) are active at this time.Abbreviations CAM
Crassulacean acid metabolism
- GCMS
gas chromatography-mass spectrometry
- MS
mass spectrometry
- NMR
nuclear magnetic resonance spectrometry
- PEP
phosphoenolpyruvate
- RuBP
ribulose 1,5-bisphosphate 相似文献
7.
Analogues of dimyristoylphosphatidylethanolamine (DMPE) have been prepared with head groups modified by N-alkylation, alkylation of carbon 2 of the ethanolamine group, or interposition of extra methylene segments between the phosphoryl and amino groups. The phases formed by these lipids in aqueous dispersions have been examined by high-sensitivity differential scanning calorimetry and Raman spectroscopy. All of the DMPE analogues examined, excepting N-methyl-DMPE but including N-ethyl-DMPE, form hydrated gel phases that are metastable with respect to a dehydrated "high-melting" solid phase that has been observed previously for DMPE itself. The properties and the conditions of formation of this high-melting phase are qualitatively distinct from those of the "subgel" phase, which is observed for dipalmitoylphosphatidylcholine and for some of the DMPE analogues examined in this study. The high-melting phases of different DMPE analogues all exhibit similarly tight packing of the acyl chains, which however do not pack according to a single type of subcell that can be universally and specifically associated with this phase. Increasing the size of the PE head group invariably decreases the melting temperature of the hydrated gel phase, even when the normal hydrogen-bonding capability of the head group is preserved. By contrast, addition of larger alkyl substituents to either the amino group or carbon 2 of the ethanolamine moiety substantially increases the transition temperature of the high-melting solid phase, indicating that the contributions of the head group to the energies of the hydrated gel and the high-melting phases are fundamentally different. Our results suggest that the head group structural requirements for a neutral phospholipid to form stable hydrated bilayers are rather stringent, a fact that may explain the overwhelming predominance of only a few such head group structures in most natural membranes. 相似文献
8.
Carbon isotope effect on dehydration of bicarbonate ion catalyzed by carbonic anhydrase 总被引:5,自引:0,他引:5
The carbon-13 kinetic isotope effect on the dehydration of HCO3- by bovine carbonic anhydrase has been measured. To accomplish this, bicarbonate was added to a buffer solution at pH 8 containing carbonic anhydrase under conditions where purging of the product CO2 from the solution is rapid. Measurement of the isotopic composition of the purged CO2 as a function of the concentration of carbonic anhydrase permits calculation of the isotope effect on the enzymic reaction. The isotope effect on the dehydration is k12/k13 = 1.0101 +/- 0.0004. This effect is most consistent with a ping-pong mechanism for carbonic anhydrase action, in which proton transfer to or from the enzyme occurs in a step separate from the dehydration step. Substrate and product dissociation steps are at least 2-3-fold faster than the hydration/dehydration step. 相似文献
9.
William M. O'Leary 《Journal of bacteriology》1959,78(5):709-713
10.
Immunocytochemical localization of the major glutathione S-transferases in adult Schistosoma mansoni
Indirect immunofluorescence was used to investigate the tissue distribution of the major isoenzymes of Schistosoma mansoni glutathione S-transferase (GSH S-transferase). When polyclonal rabbit antisera against GSH S-transferase isoenzymes SmGST-1, -02, and -3 were applied to cryostat or plastic-embedded sections of fixed adult worms, a punctate pattern of enzyme distribution was observed that was restricted to the parenchyma. Labeling was much more pronounced in males than females, consistent with the biochemically determined distribution of these enzymes between the sexes. Intense immunolabeling was noted within the subectocytoplasmic core tissue of the tubercles of the male that appeared to be connected to deep parenchymal cells by immunoreactive cell processes. Immunofluorescence could be blocked completely by prior incubation of antisera with affinity-purified enzyme. Although schistosome GSH S-transferases have been reported to be protective antigens, no immunoreactivity was detected within or on the tegument, including the dorsal spines of the male. The lack of tegumental immunoreactivity was confirmed by immunoblotting of tegumental membrane preparations following SDS-PAGE. Muscle fibers, vitelline cells, and cecal epithelium also failed to react. The fact that the GSH S-transferases were not uniformly distributed among all parenchymal cells suggests the existence of subpopulations of parenchymal cells that are preferentially involved in the conjugation of electrophiles with glutathione. 相似文献