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1.
Frank M. Sulzman Leland N. Edmunds Jr. 《Biochimica et Biophysica Acta (BBA)/General Subjects》1973,320(3)
Possible factors that could generate the circadian oscillations in alanine dehydrogenase (EC 1.4.1.1.) activity observed in cultures of non-dividing Euglena gracilis (Z) have been examined in an effort to learn more about the basic timekeeping mechanism of biological clocks. No differences in Km, pH optimum or electrophoretic mobility could be demonstrated between enzyme extracted from the minimum part of the 24-h oscillation in activity and that extracted from the maximum part. Also, no evidence for the presence of activators or inhibitors was found in mixing experiments. The effect of cycloheximide on the rhythm was examined; it was shown that the oscillation ceases in the presence of the inhibitor, but that if the inhibitor is removed after 12 h, the rhythm resumes with no apparent change in phase. Analyses of gel scans of enzyme preparations partially purified by (NH4)2SO4 fractionation and polyacrylamide gel electrophoresis indicated that there was more alanine dehydrogenase protein present at the maximum part of the cycle than there was at the minimum part. In view of these and other data, an operational model of a circadian biological clock is discussed. 相似文献
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F H Wolfe S K Sathe D E Goll W C Kleese T Edmunds S M Duperret 《Biochimica et biophysica acta》1989,998(3):236-250
Chicken breast muscle has three Ca2+-dependent proteinases, two requiring millimolar Ca2+ (m-calpain and high m-calpain) and one requiring micromolar Ca2+ (mu-calpain). High m-calpain co-purifies with mu-calpain through successive DEAE-cellulose (steep gradient), phenyl-Sepharose, octylamine agarose, and Sephacryl S-300 columns, but elutes after mu-calpain when using a shallow KCl gradient to elute a DEAE-cellulose column. The mu- and m-calpains have 80 and 28 kDa polypeptides and are analogous to the mu- and m-calpains that have been purified from bovine, porcine and rabbit skeletal muscle. High m-calpain, which seems to be a new Ca2+-dependent proteinase, is still heterogeneous after the DEAE-cellulose column eluted with a shallow KCl gradient. Additional purification through two successive HPLC-DEAE columns and one HPLC-SW-4000 gel permeation column produces a fraction having six major polypeptides and 6-8 minor polypeptides on SDS-PAGE. A 74-76 kDa polypeptide in this fraction reacts in Western blots with monospecific, polyclonal anti-calpain antibodies that react with both the 80 kDa and the 28 kDa polypeptides of mu- or m-calpain. High m-calpain also is related to mu- and m-calpain in that it causes the same limited digestion of skeletal muscle myofibrils, has a similar pH optimum near pH 7.9-8.4, requires Ca2+ for activity, and reacts with the calpain inhibitor, calpastatin, and a variety of serine and cysteine proteinase inhibitors in a manner identical to mu- and m-calpain. High m-calpain differs from mu- and m-calpain in its elution off DEAE-cellulose columns and its requirement of 3800 microM Ca2+ for one-half maximal activity compared with 5.35 microM Ca2+ for mu-calpain and 420 microM Ca2+ for m-calpain. The physiological significance of high m-calpain in unclear. The presence of mu-calpain in chicken breast muscle suggests that all skeletal muscles contain both mu- and m-calpain, although the relative proportions of these two proteinases may vary in different species. 相似文献
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Dr. Isabelle A. Carré Ady S. Oster Danielle L. Laval-Martin Leland N. Edmunds Jr. 《Current microbiology》1989,19(4):223-229
The photosynthesis-deficient ZC mutant ofEuglena gracilis Klebs (strain Z) was cultured at 16°C on an aerated, magnetically stirred, mineral medium containing 0.1% ethanol (pH 7.0). Cell division could be entrained by a 12: 12 light: dark cycle (LD: 12, 12) or even by a one-pulse skeleton photoperiod (LD: 1,23) The rhythm free-ran in DD for at least 8 days with a circadian period (=25.5 h) in populations that had been previously entrained by LD. The freerunning rhythm could be phase-shifted by a single 1-h light pulse (3000 lx). The strong (Type 0) phase-response curve derived from the resetting effects of such signals given at different circadian times was similar to that for the photosynthetic wild-type strain. These results demonstrate that the presence of a functional chloroplast compartment is not necessary for the circadian clock to function inEuglena and suggest that phase resetting of the circadian clock by light occurs via a similar pathway in both photosynthetic and non-photosynthetic cell types. 相似文献
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D L Laval-Martin I A Carré S J Barbera L N Edmunds 《Archives of biochemistry and biophysics》1990,276(2):433-441
NAD kinase and NADP phosphatase activities were detected in the supernatant and the pellet fractions prepared by sonication and centrifugation of the achlorophyllous ZC mutant of the phytoflagellate Euglena gracilis. A detailed study of substrate concentration-velocity curves enabled us to define the saturating substrate concentrations that were used in the enzyme assays. An analysis of the reproducibility of the entire assay procedure indicated that the pooled standard error was about 14%. We report circadian variations in the activities of NAD kinase and NADP phosphatase in the soluble and membrane-bound fractions of both synchronously dividing and nondividing cultures maintained in constant darkness. Bimodal circadian rhythms in total NADP phosphatase activity were found in dividing cells (peaks at circadian times [CT] 00 and 12). The peak observed at CT 00-03 disappeared when the cells had ceased dividing, a result that suggests that it might be regulated by the cell division cycle. NAD kinase activity displayed unimodal circadian rhythms (peak at CT 12) in dividing cells, which persisted with the same phase after the culture entered the stationary phase of growth. Results are discussed with reference to a model (K. Goto, D. L. Laval-Martin, and L. N. Edmunds, Jr., 1985, Science 228, 1284-1288) in which we have proposed that the Ca2(+)-transport system, Ca2+, calmodulin, NAD kinase, and NADP phosphatase could represent clock "gears" that might constitute a self-sustained circadian oscillating loop. 相似文献
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Cell division cycles and circadian clocks : phase-response curves for light perturbations in synchronous cultures of euglena 总被引:1,自引:1,他引:0 下载免费PDF全文
The cell division rhythm in Euglena gracilis Klebs (Z strain) freeruns with a circadian period (30.2 ± 1.8 hours for 156 monitored oscillations) in aerated, magnetically stirred, 8-liter, axenic batch cultures grown photoautotrophically at 25°C in LD: 3,3, (7,500 lux, cool-white fluorescent) 6-hour light cycles from the moment of inoculation. Cell number was measured at 2-hour intervals with an automatic fraction collector and Coulter Electronic Particle Counter. At different circadian times throughout the 30-hour division cycle, 3-hour light perturbations were imposed on free-running cell populations by giving light during one of the intervals when dark would have fallen in the LD: 3,3 regimen. Using the onset of division as the phase reference point, the net steady-state phase advance or delay (±Δ) of the rhythm was determined after transients, if any, had subsided (usually in one or two days) relative to an unperturbed control culture. Both +Δ and −Δ were found, with maximum values of approximately ±11 to 12 hours being obtained at circadian time (CT) 20 to 22 (the `breakpoint'); little, if any phase shift occurred if the light signal was given between CT 6 and CT 12. The phase-resetting curve obtained by plotting new phase (′) versus old phase () was of the type 0 (`strong') variety. Light perturbations, no matter when imposed, engendered new phases which mapped to a relatively restricted portion (CT 6 to CT 13) of the circadian cycle.
These data provide the first detailed phase-response curve for a circadian mitotic clock. The findings, therefore, not only further support the hypothesis that a circadian oscillator (perhaps exhibiting limit cycle behavior) can modulate cell division in eukaryotic cells, but also provide a useful basis for the dissection of the nature and extent of the coupling between cell division and circadian cycles.
相似文献8.
For nearly 50 years, analyses of coral physiology have used small coral fragments (nubbins) to make inferences about larger colonies. However, scaling in corals shows that linear extrapolations from nubbins to whole colonies can be misleading, because polyps in nubbins are divorced of their morphologically complex and physiologically integrated corallum. We tested for the effects of integration among branches in determining size-dependent calcification of the coral Pocillopora spp. under elevated PCO2. Area-normalized net calcification was compared between branches (nubbins), aggregates of nubbins (complex morphologies without integration) and whole colonies (physiologically integrated) at 400 versus approximately 1000 µatm PCO2. Net calcification was unaffected by PCO2, but differed among colony types. Single nubbins grew faster than whole colonies, but when aggregated, nubbins changed calcification to match whole colonies even though they lacked integration among branches. Corallum morphology causes the phenotype of branching corals to differ from the summation of their branches. 相似文献
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Mattias Ljunggren Karin Willquist Guido Zacchi Ed WJ van Niel 《Biotechnology for biofuels》2011,4(1):1-15