首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献,搜索用时 15 毫秒
1.
Abstract— The kinetics of phytochrome phototransformation from the red-absorbing form (Pr) to the far-red-absorbing form (Pfr) in vivo at 22°C were studied using a double flash apparatus with 1-ms flashes. Photoconversion by simultaneous flashes of red light saturates at a low Pfr level, indicating the possible attainment of a photoequilibrium between the excitation of Pr and the photoreversion of intermediates in the course of the I-ms flashes. At saturation energy, simultaneous flashes resulted in about 50% as much Pfr as was produced by saturating irradiation with 5 s red light. Intermediates of the phototransformation pathway were analysed by separating two red or a red and a far-red flash by variable dark intervals. In both plants phototransformation intermediates with half-lives < 1 ms occur, but they are too short-lived to characterize by our method. The subsequent intermediates have half-lives of about 7 ms and 150 ms in A vena , 2 ms and 10 ms in Mougeotia. The conversion from Pr to Pfr seems to be completed 1 s after the red flash in Avena. In the alga Mougeotia , Pfr formation seems to be finished within only 50 ms after the inducing red flash. The kinetics obtained from physiological and spectrophotometric experiments with Avena mesocotyls are almost identical. These observations indicate that the physiological response corresponds directly to the amount of Pfr produced and not to phototransformation intermediates or "cycling" between Pr and Pfr.  相似文献   

2.
Abstract— The phytochrome-encoding gene Cerpu;PHY;2 ( CP2 ) of the moss Ceratodon purpureus was heterologously expressed in Saccharomyces cerevisiae as a polyhistidine-tagged apoprotein and assembled with phytochromobilin (P φ B) and phycocyanobilin (PCB). Nickel-affinity chromatography yielded a protein fraction containing approximately 80% phytochrome. The holoproteins showed photoreversibility with both chromophores. Difference spectra gave maxima at 644/716 nm (red-absorbing phytochrome [Pr]) far-red-absorbing phytochrome [Pfr]) for the PCB adduct, and 659/724 nm for the PφB-adduct, the latter in close agreement with values for phytochrome extracted from Ceratodon itself, implying that PφB is the native chromophore in this moss species. Immunoblots stained with the antiphytochrome antibody APC1 showed that the recombinant phytochrome had the same molecular size as phytochrome from Ceratodon extracts. Further, the mobility of recombinant CP2 holophyto-chrome on native size-exclusion chromatography was similar to that of native oat phytochrome, implying that CP2 forms a dimer. Kinetics of absorbance changes during the Pr→ Pfr photoconversion of the PCB adduct, monitored between 620 and 740 nm in the microsecond range, revealed the rapid formation of a red-shifted intermediate (I70o)> decaying with a time constant of - 110 u.s. This is similar to the behavior of phytochromes from higher plants when assembled with the same chromophore. When following the formation of the Pfr state, two major processes were identified (with time constants of 3 and 18 ms) that are followed by slow reactions in the range of 166 ms and 8 s, respectively, albeit with very small amplitudes.  相似文献   

3.
Abstract— The phototransformation of native (124 kDa)oat phytochrome, Pr Pfr, Has been studied at 10C by two laser/ two-color flash photolysis. the overall PrPfr reaction yield did not vary with temperature within the range4–21C. Foloeing the excitation of Pr with a single 15 ns laser flash at 650nm, the formation of Pfr was quantitavely measured in a time-resolved experiment in the presence of a second 8 ns laser flash at 710 nm delayed from the initial flash. the second laser flash causes at 1.0 s after the initial laser flash a depletion of the uintermediate I700 as welll as a reduction of the Pfr absorption at 730 nm. The depletion of I700 correlates quantitavely with the reduction of Pfr formation. The absorpton spectra of I700 and of the following intermendiate, Ibi, were calculated assuming that the amount of Pr, which is photoconverted by a single laser, equals the amount of Pfr formed.  相似文献   

4.
Abstract— Irradiation of small phytochrome from oat in its Pr form with 15 ns laser pulses of different wavelengths(605–655 nm) gave rise to a difference absorption with maxima at 400 and 685 nm for the first detectable transient. Bleaching of a 660 nm band was observed, non-recuperable up to 1 ms. The transient absorption has a lifetime of 70±15 μs at 273 K. The transient is tentatively identified as lumi-R and the conformation of its chromophore is postulated to be more extended than that of Pr. A deviation from the exponential decay of the lumi-R absorption at 284 and 300 K and the lack of observable enhancement of the far-red absorption within 1 ms are interpreted in terms of the appearance of still other intermediates on this time scale between lumi-R and Pfr phytochrome.  相似文献   

5.
Abstract— The circular dichroism spectra of oat phytochrome were recorded. Qualitatively, the same spectra were found for large (360 kilodaltons) and small (60 kilodaltons) phytochrome. Quantitative CD data were reported for small Pr and Pfr (photoequilibrium mixture with 20% Pr) in tris buffer (native state) and in acid urea (denatured state). Further, the CD spectra of a phytochromobilinpeptide in acid solution with and without urea were recorded. Differences between the data in native and denatured state are discussed.  相似文献   

6.
Abstract. Phototransformation kinetics of 124-kDa oat phytochrome at 298 K after a red (660-nm) laser flash excitation were recorded at different wavelengths. The kinetics of the dark relaxation processes for lumi-R to Pfr can be satisfactorily described by only 3 rate constants: k = 28000 s-1 370 s-1 and 20 s-1. The first rate constant is due to the decay of lumi-R to meta -Ra. The latter two rate constants correspond to processes establishing the far-red (>700 nm) absorption band. No meta -Rb could be detected. From the wavelength dependency of the amplitudes of these two rates, parallel pathways in the formation of Pfr could be excluded. A unique sequential pathway for the dark relaxation leading to Pfr seems to be an intrinsic property of 124-kDa phytochrome, however. Assuming a sequential pathway, molar extinction coefficients for intermediates have been calculated. These values agree with molar extinction coefficients obtained from low-temperature spectra. The process with a rate constant of 370 s-1 corresponds to absorbance changes for the formation of meta -Rc from meta -Ra and the rate constant of 20 s-1 describes the absorbance changes due to the transformation of meta -Rc to Pfr.  相似文献   

7.
Abstract— In epicotyl tissue of Pisum , irradiation of Pr at – 196°C forms a stable product P698, whereas Pfr forms a stable product P650. On warming P698, dark transformation to Pr predominates. On warming P650 to – 70°C an intermediate P690 is formed which bleaches on further warming to –10°C. When tissue is cooled to –196°C during actinic irradiation, difference spectra for subsequent warming to –10°C indicate that Pr, Pfr and an intermediate P710 are formed from weakly absorbing intermediates. Complete photoconversion of Pr to Pfr is not possible at temperatures below –5°C. As the temperature is reduced, the amount of Pfr produced from Pr decreases, while P710 increases. P710 can be photoconverted at –20°C and above, ultimately forming Pr, but in contrast to Pfr it is not photoconvertible at –196°C.  相似文献   

8.
In the blue spectral region, the phototransformation difference spectrum of oat phytochrome extracted as Pfr differs from that of phytochrome extracted as Pr. The difference absorbance maximum for phytochrome extracted as Pfr is at 420 nm, while that extracted as Pr is at 412 nm. The phototransformation difference spectrum measured in the blue in oat coleoptile tips without inner leaves, corresponds very well with that of phytochrome as extracted in its Pfr form. There is, however, a slight apparent attenuation of the blue difference band relative to those in the red-far-red. In coleoptile tissue containing inner leaves, the blue difference band is relatively even more highly attenuated. A similar attenuation is observed in the blue, in the protochlorophyllide to chlorophyllide phototransformation difference spectrum. In the spectrum measured with excised coleoptile without inner leaves, there is a small attenuation, while in coleptile tissue with inner leaves the attentuation is nearly 9-fold. These data suggest that the observed attenuation is probably artifactual. Neither instrumental non-linearity nor fluorescence induced by the measuring beam could explain the observed attenuation. It is suggested that the observed attenuation is probably mainly the result of wavelength dependent scatter amplification, the amplification in the blue being attenuated by the high background absorption of other pigments in this region.  相似文献   

9.
Abstract— Irradiation of the Pr form of phytochrome in the presence of flavin mononucleotide (FMN) which absorbs the actinic blue light yields Pfr at a rate greater than that in the absence of FMN. The actinic blue light absorbed by FMN enhances the phototransformation of Pr via the energy transfer from the former to the latter. On the other hand, the photoreversion of Pfr was inhibited by the presence of FMN when illuminated with blue light. The lack of photo-enhancement of the reversion of Pr, by blue light suggests that the Pfr chromophore (acceptor) transition dipole is virtually perpendicular to the FMN transition dipole, as the result of a chromophore reorientation in the Pr→Pfr phototransformation. The fact that blue light absorbed by flavin preferentially enhances the forward phototransformation of phytochrome while inhibiting the reversion may have an important implication in the high irradiance responses in plants in terms of a preferential accumulation of Pfr by blue light excitation.  相似文献   

10.
Abstract— Protonemata of the moss Ceratodon purpureus cultured in white light were transferred to darkness for 3 days and then used for phototropic experiments. Irradiation of the apical region of vertically position protonemata with small beams (0.2 mm) of red light induced a growth response towards the irradiated side (positive phototropism). The phototropic response showed irradiance dependence. The effect of red light was completely reversed by far-red light following red light irradiations, demonstrating that phytochrome was the photoreceptor pigment. Far-red light or UV-blue light had no influence on either bulging or phototropism. Experiments with linearly polarized red or far-red light showed a different dichroic distribution of phytochrome in its different forms, the red-absorbing form, Pr and the far-red-absorbing form, Pfr. Red light with a vibration plane parallel to the long axis of the filaments was most effective. The effectiveness of far-red light was expressed best when its vibration plane was 90° to the electrical vector of the inductive red light.  相似文献   

11.
Abstract— The time courses for Pr appearance, Pr disappearance and Pfr destruction have been analysed in cotyledons of Cucurbita pepo L. after different preirradiation programs. In etiolated seedlings the rate of Pr appearance is low in young seedlings reaching a maximum in 3.5–5 day old seedlings then decreasing rapidly with increasing age. The rate of Pfr destruction is very low in young seedlings, increases rapidly up to the 4th day and then remains almost constant. The disappearance of Pr becomes significant for seedlings older than 45 days. These reactions seem not to be influenced by short preirradiations. However, after prolonged preirradiation, a degree of control of P, appearance and/or disappearance by the "internal clock appears to be operative.  相似文献   

12.
Abstract— The changes in the chromophore structure of pea phytochrome during phototransformation in vitro from the red-light-absorbing form (Pr) to the far-red-light-absorbing form (Pfr), and from Pfr to Pr, were analysed in terms of wavelength and oscillator strength of absorption, using the zero-differential overlap approximation of the molecular orbital theory for electrons. The effect of a point-charge and a point-dipole on the optical absorption of phytochromobilin intermediates were examined using the stationary perturbation theory for degenerate states. The results indicate that the cis-trans photoisomerization of the pyrrole ring D, if any, occurred within 10 μs after a laser-flash excitation of the phytochrome, and that the conformations of phytochromobilin and the protein moiety of phytochrome were not significantly changed during the period of examination of phototransformation in either direction.  相似文献   

13.
Abstract— Kinetic experiments have provided evidence for a series of light and dark reactions of phytochrome intermediates at low temperature in Pisum epicotyl tissue. A photoequilibrium exists between Pr and P698, and between Pfr and P650. A dark reversion of P698Pr and P650pfr at –70°C has been demonstrated. When cooled to 70°C under incandescent light, most of the phytochrome in the tissue is driven into photochemically unreactive intermediates. About 2% of the phytochrome remains as weakly absorbing intermediates that form Pr and Pfr in darkness. A scheme is presented for phytochrome phototransformation in vivo.  相似文献   

14.
DEPENDENCE OF Pfr/Ptot-RATIOS ON LIGHT QUALITY and LIGHT QUANTITY   总被引:2,自引:0,他引:2  
Abstract— Not only the spectral distribution of the light source determines the relative proportion of phytochrome in the Pfr(Pr) form, the Pfr/Ptot-ratio also depends strongly on the fluence rate of the irradiation. This dependence has been observed in the cotyledons of etiolated mustard seedlings for blue light of fluence rates below 20 Wm-2. It has also been observed for white light and seems to be a characteristic of the phytochrome system resulting from the involvement of phytochrome thermal reactions as well as Pr Pfr photoconversions. The fluence rate dependence of Pfr/Ptot-ratios can be used to analyze the characteristic transformations of the phytochrome system. Phototransformations together with a fast thermal transformation (τ½⋍ 3min) are consistent with the results obtained for blue and white light.  相似文献   

15.
RESONANCE RAMAN SPECTRA OF THE Pr-FORM OF PHYTOCHROME   总被引:1,自引:0,他引:1  
Abstract— Resonance Raman spectra of the Pr-form of oat phytochrome have been obtained at 77 K. Interference from phytochrome fluorescence is avoided by employing far-red 752 nm excitation. Vibrational assignments are suggested for the tetrapyrrole chromophore in phytochrome by comparison with previously published model compound spectra and by examining the characteristic shifts induced by deuteration of the pyrrole nitrogens. The lack of carbonyl intensity, the frequencies of the 1626 and 1644 cm-1 C=C stretching modes, and the presence of an intense mode at 1326 cm-1 are all consistent with a protonated structure for the tetrapyrrole chromophore in Pr. This suggests that the -50 nm red-shift of the protein-bound chromophore absorption compared to the chromophore in vitro is caused by protonation of the pyrrole nitrogen.  相似文献   

16.
The dichroic orientation of phytochrome observed both in the phytochrome-mediated phototropism in Adiantum protonemata and in the phytochrome-mediated chloroplast movement in Mougeotia were analyzed in terms of the orientation of the transition moment associated with the long-wavelength absorption band, assuming that phytochrome, associated with the plasma membrane, rotates around the normal to the membrane. The orientation of the long-wavelength transition moment of the phytochrome chromophore was calculated using the zero-differential overlap approximation of the molecular orbital theory for ir-electrons. The results indicate that the orientation of the long-wavelength transition moment mainly changes later than 2 ms after red light excitation of Pr, and that the different dichroic orientations of Pr and Pfr can be attributed to the change in the angle of the long-wavelength transition moment of phytochrome with the plasma membrane from 18o to 72o during phototransformation.  相似文献   

17.
Abstract— Accumulation of weakly absorbing phytochrome intermediates has been demonstrated in Pisum epicotyl tissue under conditions of pigment cycling using a quasi-continuous measuring spectrophotometer. An action spectrum shows 690–700 nm to be the most efficient wavelength range in this process. Difference spectra for the decay of intermediates maintained by 690 nm light show that, if the experiment is done at 0°C, only Pfr is formed. At – 11°C, intermediates decaying to Pr can also be observed. At – 20°C, Pr is produced as well as a pigment with peak absorption at 710nm. Kinetic analysis of intermediate decay at – 11°C reveals that at least two intermediates are maintained by 690 nm light. The level of intermediate maintained by incandescent light at 0°C was 25% higher in air than in nitrogen.  相似文献   

18.
Abstract— In the cotyledons of the mustard seedling Sinapis alba L. the duration of the Shibata shift can be greatly shortened by a pretreatment with light pulses prior to the protochlorophyllide– chloro-phyllide a photoconversion. It was shown that the light pulses act through photochrome (P fr ). Since reversibility of a red light pulse induction by a far-red light pulse is rapidly lost (within 2 min) it is concluded that at least the initial action of Pfr occurs rapidly in this response. On the other hand, the effect of a red light pulse on the rate of protochlorophyll regeneration in the mustard seedling cotyledons is fully reversible by a far-red light pulse for more than 5 min. It is concluded that control of protochlorophyll regeneration and control of the Shibata shift by phytochrome cannot be consequences of the same initial action of Pfr Apparently Pfr controls both phenomena independently.  相似文献   

19.
Abstract— Single-celled protonemata of Adiantum capillus-veneris were cultured under continuous red light for 6 days and then in the dark for 15 h. Brief local exposure of a flank (5 times 20 /mi) of the subapical region of a protonema to a microbeam of red light effectively induced a phototropic response toward the irradiated side. The degree of the response was dependent upon the fluence of the red light. Red/far-red reversibility was typically observed in this photoreaction, showing that phytochrome was the photo-receptive pigment. When the flank was irradiated with a microbeam of linearly polarized red and far-red light, red light with an electrical vector parallel to the cell surface was most effective. However, the far-red light effect was most prominent when its electrical vector was normal to the cell surface. These polarized light effects indicate the different dichroic orientation of Pr (red-light-absorbing form of phytochrome) and Pr (far-red-light-absorbing form of phytochrome) at the cell flank.  相似文献   

20.
-Large phytochrome immobilized via anti-phytochrome immunoglobulin bound to Sepharose beads was irradiated to saturation with unpolarized far-red light. The apparent absorbance level was recorded in a dual wavelength spectrophotometer with both measuring beams set to either 660 or 730 nm and polarized perpendicular to each other. The sample was then irradiated with red polarized light. The apparent change in absorbance obtained after this irradiation indicated that purified phytochrome could show linear dichroism. From the absorbance values obtained it was computed that the direction of the long-wavelength transition moment changes by either 32 or 148o, when phytochrome is transformed from Pr to Pfr. Considering the model of Hahn and Song (1981) the latter value appears more likely. In light of these results, the conclusions drawn from in vivo experiments on action dichroism in Dryopteris (Etzold, 1965), Adiantum (Kadota et al., 1982) and Mougeoutia (Haupt. 1970), which point to a 90o rotation. should be reconsidered.  相似文献   

设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号