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1.
The nanosecond fluorescence depolarization method was applied to measure the fluorescence lifetime () and the rotational correlation time () of bovine serum albumin (BSA) labeled with 1-dimethylaminonaphthalene-5-sulfonyl chloride (dansyl-Cl). Changes of and of dansyl BSA in the guanidine denaturation and in the thermal denaturation were examined. In parallel, the secondary structural change of dansyl BSA was followed by circular dichroism measurements. The magnitude of was almost unchanged between 1 and 2 M guanidine, where the secondary structure of the protein was predominantly disrupted; whereas that of began to increase before the disruption of secondary structure in the guanidine denaturation. In the thermal denaturation, in contrast, changes of both and occurred in a temperature range where the secondary structure was predominantly disrupted. The volume of equivalent sphere (V
e
) and the axial ratio () for the BSA were 3.6–3.8×10–19 cm3 and 3.6 at 2M guanidine as against 2.1×10–19 cm3 and 2.2 in the absence of guanidine (25°C), respectively. The magnitudes ofV
e
and were 4.9×10–19 cm3 and 4.5 at 65°C, respectively. Although the secondary structural change of dansyl BSA was irreversible in the thermal denaturation,V
e
and were reversible. 相似文献
2.
The helicities in various fragments of bovine serum albumin (BSA) were examined in the thermal denaturation and in sodium docecyl sulfate (SDS) denaturation. The thermal denaturation was examined in a temperature range between 2 and 65°C. The helicity decreased with a rise of temperature and it recovered to some degree upon cooling temperature. A rather high reversibility was observed in the BSA fragments, which were located in the N-terminal of the parent protein and then contained the first large loop with no disulfide bridge. The high reversibility was available also for the helicity in the first large loop of the fragment, disulfide bridges of which were reduced. The fragments, which were smaller than one domain, became unstable in the SDS denaturation. The helicities of such fragments decreased in lower SDS concentrations compared with those of the intact BSA and the large fragments, which contained one or more domains. A resistance to the SDS denaturation appeared in the helices of every large loop even after the fragmentation. On the other hand, helicities of the fragments decreased to 20–25% upon the reduction of disulfide bridges. However, the helicities of these fragments increased to 35–40% in the SDS denaturation. 相似文献
3.
Jakub Psencík Geoffrey F. W. Searle Jan Hála Tjeerd J. Schaafsma 《Photosynthesis research》1994,40(1):1-10
Fluorescence Detected Magnetic Resonance (FDMR) spectra have been measured for whole cells and isolated chlorosomal fractions for the green photosyntheic bacteria Chlorobium phaeobacteroides (containing bacteriochlorophyll e, and isorenieratene as major carotenoid) and Chlorobium limicola (containing bacteriochlorophyll c, and chlorobactene as major carotenoid). The observed transition at 237 MHz (identical in both bacteria) and > 1100 MHz can be assigned, by analogy with published data on other carotenoids, to the 2E and D + E transitions, respectively, of Chlorobium carotenoids. Their zero field splitting (ZFS) parameters are estimated to be: |D|=0.0332 cm–1 and |E|=0.0039 cm–1 (chlorobactene), and |D|=0.0355 cm–1 and |E|=0.0039 cm–1 (isorenieratene). In the intermediate frequency range 300–1000 MHz the observed transitions can be assigned to chlorosomal bacteriochlorophylls c and e, and to bacteriochlorophyll a located in the chlorosome envelope and water-soluble protein. The bacteriochlorophyll e triplet state measured in 750 nm fluorescence (aggregated chlorosomal BChl e) is characterised by the ZFS parameters: |D|=0.0251 cm–1 and |E|=0.0050 cm–1.Abbreviations BChl -
bacteriochlorophyll
- BPh -
bacteriopheophytin
-
Chl. -
Chlorobium
- F(A)(O)DMR -
fluorescence (absorption) (optical) detected magnetic resonance
- FF -
fluorescence fading
- ISC -
intramolecular intersystem crossing
- RC -
reaction center
- ZFS -
zero field splitting 相似文献
4.
Summary The utility of the lipophilic anion thiocyanate (SCN+) as a probe for the indirect estimation of the cell membrane potential (V
m
) in Ehrlich ascites tumor cells has been evaluated by comparison to direct electrophysiological measurements. SCN accumulation is consisten with first-order uptake into a single kinetically-identifiable cellular compartement, achieving steadystate distribution in 20–30 min at 22°C. The steady-state distribution ratio ([SCN–]
c
/[SCN–]
e
) in physiological saline is 0.44±0.02. Treatment of the cells with proparanolol (0.13 mM), an activator of Ca2+ dependent K+ channels, reduces the steady-state distribution ratio to 0.19±0.02. Conversely, treatmetn with BACl2 (10 mM), an antagonist of the pathway, increases the SCN– distribution ratio to 0.62±0.01. The equilibrium potentials (V
SCN
) calculated under these conditions are virtually identical to direct electrophysiological measurements of theV
m
made under the same conditions. The effect of varing extracellular [K+]([K+]
e
) in the presence of constant [Na+]
e
=100 mM has also been tested. In control cells, elevation of [K+]
e
from 6 to 60 mM reducesV
SCN
from –20.6±1.0 to –13.2±1.2 mV. Again, microelectrode measurements give excellent quantitative agreement. Propranolol increases the sensitivity of the cells to varying [K+]
e
, so that a 10-fold elevation reducesV
SCN
by approximately 31 mV. BaCl2 greatly reduces this reponse: a 10-fold elevation in [K+]
e
yielding only a 4-mV rediction inV
SCN
. It is concluded that the membrane potential of Ehrlich cells can be estimated accurately from SCN– distribution measurements. 相似文献
5.
The sexual agglutinin from the mating-type minus gametes ofChlamydomonas reinhardtii was purified by gel filtration and hydroxyapatite chromotography. The minus agglutinin was identified as a single glycopolypeptide termed Agg(-) of very high molecular weight by SDS-poly-acrylamide gel electrophoresis. It was also observed as a glycoprotein with agglutinin activity on non-SDS polyacrylamide gels. The native agglutinin appeared to be composed of a complex of Agg(-) subunits. It consisted of about 60% protein and 40% carbohydrate and the activity was diminished by a mild periodate oxidation. When the plus agglutinin was also purified and compared with the minus agglutinin, it was found that both agglutinins migrate in the same position by SDS-polyacrylamide gel electrophoresis, whereas their behaviors on gel filtration and hydroxyapatite chromatography are different.Abbreviations
mt
+/–
mating-tape plus or minus
- SDS
sodium dodecyl sulfate
- Ve
elution volume
- Vo
void volume
- kDa
kilodalton 相似文献
6.
M Vitolo M A Duranti M B Pellegrim 《Journal of industrial microbiology & biotechnology》1995,15(2):75-79
S. cerevisiae was grown in a blackstrap molasses containing medium in batch and fed-batch cultures. The following parameters were varied: pH (from 4.0 to 6.5), dissolved oxygen (DO) (from 0 to 5.0 mg O2L–1) and sucrose feeding rate. When glucose concentration (S) was higher than 0.5 g L–1 a reduction in the specific invertase activity of intact cells (v) and an oscillatory behavior of v values during fermentation were observed. Both the invertase reduction and the oscillatory behavior of v values could be related to the glucose inhibitory effect on invertase biosynthesis. The best culture conditions for attainingS. cerevisiae cells suitable for invertase production were: temperature=30°C; pH=5.0; DO=3.3 mg O2L–1; (S)=0.5 g L–1 and sucrose added into the fermenter according to the equations: (V–Vo)=t2/16 or (V–Vo)=(Vf–Vo)·(e0.6t–1)/10.This work was supported by FAPESP 相似文献
7.
Estimation of the effect of photoinhibition on the carbon gain in leaves of a willow canopy 总被引:21,自引:0,他引:21
The occurrence of photoinhibition of photosynthesis in leaves of a willow canopy was examined by measuring the chlorophyll-a fluorescence ratio of F
V/F
M (FM is the maximum fluorescence level of the induction curve, and FV is the variable fluorescence, F
V=F
M–F
0, where F0 is the minimal fluorescence). The majority of the leaves situated on the upper parts of peripheral shoots showed an afternoon inhibition of this ratio on clear days. This was the consequence of both a decrease in F
M and a rise in F
O. In the same leaves the diurnal variation in intercepted photosynthetic photon flux density (PPFD) was monitored using leaf-mounted sensors. Using the multivariate method, partial least squares in latent variables, it is shown that the dose of PPFD, integrated and linearly weighted over the last 6-h period, best predicts photoinhibition. Photoinhibition occurred even among leaves that did not intercept PPFDs above 1000 mol·m–2·s–1. Exposure of leaves to a standard photoinhibitory treatment demonstrated that the depression in the F
V/F
M ratio was paralleled by an equal depression in the maximal quantum yield of CO2 uptake and a nearly equal depression in the rate of bending (convexity) of the light-response curve of CO2 uptake. As a result, the rate of net photosynthesis is depressed over the whole natural range of PPFD. By simulating the daily course in the rate of net photosynthesis, it is estimated that in the order of one-tenth of the potential carbon gain of peripheral willow shoots is lost on clear days as a result of photoinhibition. This applies to conditions of optimal temperatures. Photoinhibition is even more pronounced at air temperatures below 23° C, as judged from measurements of the FV/FM ratio on clear days: the afternoon inhibition of this ratio increased in a curvilinear manner from 15% to 25% with a temperature decrease from 23° to 14° C.Abbreviations and Symbols FO
minimum fluorescence
- FV
variable fluorescence
- FM
maximum fluorescence
- PLS
partial least squares in latent variables
- PPFD
photosynthetic photon flux density
- VPD
water vapour-pressure deficit
This study was supported by the Swedish Natural Science Research Council. We are indebted to Dr. Jerry Leverenz (Department of Plant Physiology, University of Umeå, Sweden) for guidance with the modelling of the photosynthesis data. 相似文献
8.
Summary Cellular impalements were used in combination with standard transepithelial electrical measurements to evaluate some of the determinants of the spontaneous lumen-positive voltage,V
e
, which attends net Cl– absorption,J
Cl
net
, and to assess how ADH might augment bothJ
Cl
met
andV
e
in the mouse medullary thick ascending limb of Henle microperfusedin vitro. Substituting luminal 5mm Ba++ for 5mm K+ resulted in a tenfold increase in the apical-to-basal membrane resistance ratio,R
c
/R
bl
, and increasing luminal K+ from 5 to 50mm in the presence of luminal 10–4
m furosemide resulted in a 53-mV depolarization of apical membrane voltage,V
a
. Thus K+ accounted for at least 85% of apical membrane conductance. Either with or without ADH. 10–4
m luminal furosemide reducedV
e
andJ
Cl
net
to near zero values and hyperpolarized bothV
a
andV
bl
, the voltage across basolateral membranes; however, the depolarization ofV
bl
was greater in the presence than in the absence of hormone while the hormone had no significant effect on the depolarization ofV
a
, Thus ADH-dependent increases inV
b
were referable to greater depolarizations ofV
bl
in the presence of ADH than in the absence of ADH 68% of the furosemide-induced hyperpolarization ofV
a
was referable to a decrease in the K+ current across apical membranes, but, at a minimum, only 19% of the hyperpolarization ofV
bl
could be accounted for by a furosemide-induced reduction in basolateral membrane Cl– current. Thus an increase in intracellular Cl– activity may have contributed to the depolarization ofV
bl
during net Cl– absorption, and the intracellular Cl– activity was likely greater with ADH than without hormone. Since ADH increases apical K+ conductance and since the chemical driving force for electroneutral Na+,K+,2Cl– cotransport from lumen to cell may have been less in the presence of ADH than in the absence of hormone, the cardinal effects of ADH may have been to increase the functional number of both Ba++-sensitive conductance K+ channels and electroneutral Na+,K+,2Cl– cotransport units in apical plasma membranes. 相似文献
9.
Molecular diffusion of solutes, like sucrose in the xanthan gum fermentation, is important in order to understand the complex behavior of mass transfer mechanisms during the process. This work was focused to determine the diffusion coefficient of sucrose, a carbon source for xanthan production, using similar sucrose and xanthan concentrations to those occurring in a typical fermentation. The diaphragm cell method was used in experimental determinations. The data showed that diffusion coefficient of sucrose significantly decreases when xanthan gum concentration increases. Theoretical and semiempirical models were used to predict sucrose diffusivity in xanthan solutions. Molecular properties and rheological behavior of the system were considered in the modeling. The models tested fitted well the behavior of experimental data and that reported for oxygen in the same system.List of Symbols
A
constant in eq. (5)
-
C
pg cm–3
polymer concentration
-
D cm2 s–1
diffusivity
-
D
ABcm2 s–1
diffusivity of A through liquid solvent
-
D
APcm2 s–1
diffusivity of A in polymer solution
-
D
AWcm2 s–1
diffusivity of A in water
-
D
Pcm2 s–1
diffusivity of polymer in liquid solvent
-
E
D
gradient of the activation energy for diffusion
-
H
P
hydratation factor of the polymer in water (g of bound water/g of polymer)
-
K dyn sn cm–2
consistency index
-
K
1
constant in eq. (5)
-
K
P
overall binding coefficient [g of bound solute/cm3 of solution]/[g of free solute/cm3 of polymer free solution]
-
n
flow behavior index
-
M
Bg g mol–1
molucular weight of liquid solvent
-
M
Pg g mol–1
molecular weight of the polymer
-
M
Sg g mol–1
Molecular weight of polymer solution (= M
BXB+MPXP)
-
R cm3 atm g mol–1 K–1
ideal gas law constant
-
T K
absolute temperature
-
V
Bcm3 g mol–1
molar volume of liquid solvent
-
V
Pcm3 g mol–1
molar volume of polymer
-
V
Scm3 g mol–1
molar volume of polymer solution
-
X
B
solvent molar fraction
-
X
P
polymer molar fraction
-
polymer blockage shape factor
-
P
volume fraction of polymer in polymer solution
-
g cm–1 s–1
viscosity
-
ag cm–1 s–1
apparent viscosity of the polymer solution
-
icm3 g–1
intrinsic viscosity
-
0 g cm–1 s–1
solvent viscosity
-
Pg cm–1 s–1
polymer solution viscosity
-
R
relative viscosity (= /
0)
-
=0 g cm–1 s–1
viscosity of polymer solution obtained at zero shear rate
-
0 g cm–3
water density 相似文献
10.
Summary Determinations of plant or algal cell density (cell mass divided by volume) have rarely accounted for the extracellular matrix or shrinkage during isolation. Three techniques were used to indirectly estimate the density of intact apical cells from protonemata of the mossCeratodon purpureus. First, the volume fraction of each cell component was determined by stereology, and published values for component density were used to extrapolate to the entire cell. Second, protonemal tips were immersed in bovine serum albumin solutions of different densities, and then the equilibrium density was corrected for the mass of the cell wall. Third, apical cell protoplasts were centrifuged in low-osmolarity gradients, and values were corrected for shrinkage during protoplast isolation. Values from centrifugation (1.004 to 1.015 g/cm3) were considerably lower than from other methods (1.046 to 1.085 g/cm3). This work appears to provide the first corrected estimates of the density of any plant cell. It also documents a method for the isolation of protoplasts specifically from apical cells of protonemal filaments.Abbreviations BSA
bovine serum albumin
- ER
endoplasmic reticulum
- Vv
volume fraction 相似文献
11.
Summary Amounts and temporal changes of the release of the tracer ions K+ (86Rb+),22Na+, and36Cl– as well as of H+ in the course of action potentials inAcetabularia have been recorded. New results and model calculations confirm in quantitative terms the involvement of three major ion transport systemsX in the plasmalemma: Cl– pumps, K+ channels, and Cl– channels (which are marked in the following by the prefixes,P, K andC) with their equilibrium voltages
X
V
e
and voltage/time-dependent conductances, which can be described by the following, first approximation. Let the maximum (ohmic) conductance of each of the three populations of transporter species be about the same (P
L,
KL,C
L=1) but voltage gating be different: the pump (
p
V
e
about –200 mV) being inactivated (open,oclosed,c) at positive going transmembrane voltages,V
m; the K+ channels (K
V
e about –100 mV) are inactivated at negative goingV
m; and the Cl– channels (C
V
e: around 0 mV), which are normally closed (c) at a restingV
m (nearPVe) go through an intermediate open (o) state at more positiveV
m before they enter a third shut state (s) in series. Model calculations, in which voltage sensitivities are expressed by the factorf=exp(V
mF/(2RT)), simulate, the action potential fairly well with the following parameters (PKco10/f ks–1,PKoc1000·f ks–1,KKco200·f ks–1,Kkoc2/f ks–1,cKco500·f ks–1,CKoc5/f ks–1,CKso0.1/f ks–1,Ckos20·f ks–1). It is also shown that the charge balance for the huge transient Cl– efflux, which frequently occurs during an action potential, can be accounted for by the observation of a corresponding release of Na+. 相似文献
12.
Ashok Kumar Patel Vijay Kumar Singh Ravi Prakash Yadav A.J.G. Moir Medicherla V. Jagannadham 《Phytochemistry》2009,70(10):1210-1216
A multi-functional enzyme ICChI with chitinase/lysozyme/exochitinase activity from the latex of Ipomoea carnea subsp. fistulosa was purified to homogeneity using ammonium sulphate precipitation, hydrophobic interaction and size exclusion chromatography. The enzyme is glycosylated (14–15%), has a molecular mass of 34.94 kDa (MALDI–TOF) and an isoelectric point of pH 5.3. The enzyme is stable in pH range 5.0–9.0, 80 °C and the optimal activity is observed at pH 6.0 and 60 °C. Using p-nitrophenyl-N-acetyl-β-d-glucosaminide, the kinetic parameters Km, Vmax, Kcat and specificity constant of the enzyme were calculated as 0.5 mM, 2.5 × 10−8 mol min−1 μg enzyme−1, 29.0 s−1 and 58.0 mM−1 s−1 respectively. The extinction coefficient was estimated as 20.56 M−1 cm−1. The protein contains eight tryptophan, 20 tyrosine and six cysteine residues forming three disulfide bridges. The polyclonal antibodies raised and immunodiffusion suggests that the antigenic determinants of ICChI are unique. The first fifteen N-terminal residues G–E–I–A–I–Y–W–G–Q–N–G–G–E–G–S exhibited considerable similarity to other known chitinases. Owing to these unique properties the reported enzyme would find applications in agricultural, pharmaceutical, biomedical and biotechnological fields. 相似文献
13.
The interactions of mapenterol with bovine serum albumin (BSA) and human serum albumin (HSA) have been investigated systematically using fluorescence spectroscopy, absorption spectroscopy, circular dichroism (CD) and molecular docking techniques. Mapenterol has a strong ability to quench the intrinsic fluorescence of BSA and HSA through static quenching procedures. At 291 K, the binding constants, Ka, were 1.93 × 103 and 2.73 × 103 L/mol for mapenterol–BSA and mapenterol–HAS, respectively. Electrostatic forces and hydrophobic interactions played important roles in stabilizing the mapenterol–BSA/has complex. Using site marker competitive studies, mapenterol was found to bind at Sudlow site I on BSA/HSA. There was little effect of K+, Ca2+, Cu2+, Zn2+ and Fe3+ on the binding. The conformation of BSA/HSA was changed by mapenterol, as seen from the synchronous fluorescence spectra. The CD spectra showed that the binding of mapenterol to BSA/HSA changed the secondary structure of BSA/HSA. Molecular docking further confirmed that mapenterol could bind to Sudlow site I of BSA/HSA. According to Förster non‐radiative energy transfer theory (FRET), the distances r0 between the donor and acceptor were calculated as 3.18 and 2.75 nm for mapenterol–BSA and mapenterol–HAS, respectively. Copyright © 2015 John Wiley & Sons, Ltd. 相似文献
14.
The fluorescence quenching spectrum of bovine serum albumin (BSA) was investigated in the presence of felodipine (FLD) by spectroscopic methods including fluorescence spectroscopy and UV–Vis absorption spectroscopy. Stern–Volmer quenching was successfully applied and the corresponding thermodynamic parameters, namely enthalpy change (ΔH), free energy change (ΔG) and entropy change (ΔS) at different temperatures (304, 314 and 324 K) were calculated according to the Van't Hoff relation. This revealed that the hydrophobic interaction plays a major role in stabilizing the complex. The fluorescence spectrum of BSA was studied in presence of various concentrations of SDS surfactant. The distance (r) between donor (BSA) and acceptor (FLD) was obtained according to fluorescence resonance energy transfer (FRET). The synchronous fluorescence spectroscopy was used to investigate the effect of FLD on BSA molecule. The result shows that the conformation of BSA was changed in the presence of felodipine. Copyright © 2009 John Wiley & Sons, Ltd. 相似文献
15.
Ian Henry Lambert Else Kay Hoffmann Finn Jørgensen 《The Journal of membrane biology》1989,111(2):113-131
Summary The fluorescence intensity of the dye 1,1-dipropyloxadicarbocyanine (DiOC3-(5)) has been measured in suspensions of Ehrlich ascites tumor cells in an attempt to monitor their membrane potential (V
m
) under different ionic conditions, after treatment with cation ionophores and after hypotonic cell swelling. Calibration is performed with gramicidin in Na+-free K+/choline+ media, i.e., standard medium in which NaCl is replaced by KCl and cholineCl and where the sum of potassium and choline is kept constant at 155mm. Calibration by the valinomycin null point procedure described by Lariset al. (Laris, P.C., Pershadsingh, A., Johnstone, R.M., 1976,Biochim. Biophys. Acta
436:475–488) is shown to be valid only in the presence of the Cl–-channel blocker indacrinone (MK196). Distribution of the lipophilic anion SCN– as an indirect estimation of the membrane potential is found not to be applicable for the fast changes inV
m
reported in this paper. Incubation with DiOC3-(5) for 5 min is demenstrated to reduce the Cl– permeability by 26±5% and the NO
3
–
permeability by 15±2%, while no significant effect of the probe could be demonstrated on the K+ permeability. Values forV
m
, corrected for the inhibitory effect of the dye on the anion conductance, are estimated at –61±1 mV in isotonic standard NaCl medium, –78±3 mV in isotonic Na+-free choline medium and –46±1 mV in isotonic NaNO3 medium. The cell membrane is depolarized by addition of the K+ channel inhibitor quinine and it is hyperpolarized when the cells are suspended in Na+-free choline medium, indicating thatV
m
is generated partly by potassium and partly by sodium diffusion. Ehrlich cells have previously been shown to be more permeable to nitrate than to chloride. Substituting NO
3
–
for all cellular and extracellular Cl– leads to a depolarization of the membrane, demonstrating thatV
m
is also generated by the anions and that anions are above equilibrium. Taking the previously demonstrated single-file behavior of the K+ channels into consideration, the membrane conductances in Ehrlich cells are estimated at 10.4 S/cm2 for K+, 3.0 S/cm2 for Na+, 0.6 S/cm2 for Cl– and 8.7 S/cm2 for NO
3
–
. Addition of the Ca2+-ionophore A23187 results in net loss of KCl and a hyperpolarization of the membrane, indicating that the K+ permeability exceeds the Cl– permeability also after the addition of A23187. The K+ and Cl– conductances in A23187-treated Ehrlich cells are estimated at 134 and 30 S/cm2, respectively. The membrane potential is depolarized in hypotonically swollen cells, confirming that the increase in the Cl– permeability following hypotonic exposure exceeds the concommitant increase in the K+ permeability. In control experiments where the membrane potentialV
m
=E
K
=E
Cl
=E
Na
, it is demonstrated that cell volume changes has no significant effect on the fluorescence signal, apparently because of a large intracellular buffering capacity. The increase in the Cl– conductances is 68-fold when cells are transferred to a medium with half the osmolarity of the standard medium, as estimated from the net Cl– efflux and the change inV
m
. The concommitant increase in the K+ conductance, as estimated from the net K+ efflux, is only twofold. 相似文献
16.
Summary Cotton (Gossypium hirsutum L. var. DP 61) was grown at different temperatures during 12-h light periods, with either 1800–2000 mol photons m–2 s–1 (high photon flux density, PFD) or 1000–1100 mol m–2 s–1 (medium PFD) incident on the plants. Night temperature was 25°C in all experiments. Growth was less when leaf temperatures were below 30°C during illumination, the effect being greater in plants grown with high PFD (Winter and Königer 1991). Leaf pigment composition and the photon-use efficiency of photosynthesis were analysed to assess whether plants grown with high PFD and suboptimal temperatures experienced a higher degree of high irradiance stress during development than those grown with medium PFD. The chlorophyll content per unit area was 3–4 times less, and the content of total carotenoids about 2 times less, with the proportion of the three xanthophylls zeaxanthin + antheraxanthin + violaxanthin being greater in leaves grown at 20–21°C than in leaves grown at 33–34°C. In leaves from plants grown at 21°C and 1800–2000 mol photons m–2 s–1, zeaxanthin accounted for as much as 34% of total carotenoids in the middle of the photoperiod, the highest level recorded in this study. This finding is consistent with a protective role of zeaxanthin under conditions of excess light. At the lower temperatures, the photochemical efficiency of photosystem II, measured as the ratio of variable to maximum fluorescence yield (F
V/F
M) after 12-h dark adaptation, was 0.76 in medium PFD plants and 0.75 in high PFD plants compared with 0.83 and 0.79, respectively, at the higher temperatures. The photon-use efficiency of O2 evolution () based on absorbed light between 630 and 700nm, decreased with decrease in temperature from 0.102 to 0.07 under conditions of high PFD, but remained above 0.1 at medium PFD. Owing to compensatory reactions in these long-term growth experiments, sustained differences inF
V/F
M and were much less pronounced than the differences in chlorophyll content and dry matter, particularly in plants which had developed at high PFD and low temperature. In fact, in these plants, which exhibited pronounced photobleaching, a largely functional photosynthetic apparatus was still maintained in cells adjacent to the lower leaf surfaces. This was indicated by measurements of photon use efficiencies of photosynthetic O2 evolution with leaves illuminated first at the upper, and then at the lower surface.Abbreviations
F
O
yield of dark level fluorescence
-
F
M
maximum yield of fluorescence, induced in a pulse of saturating light
-
F
V
yield of variable fluorescence (=F
M-F
o)
- PFD
photon flux density
- iw
photon use efficiency of O2 evolution based on white (400–700 nm) incident light
- ir
photon use efficiency based on red (630–700 nm) incident light
- aw
photon use efficiency based on white absorbed light
- ar
photon use efficiency based on red absorbed light 相似文献
17.
It was found that the fluorescence of Tb3+–epinephrine (E) complex can be enhanced by both bovine serum albumin (BSA) and sodium dodecylsulfate (SDS), and stabilized by ascorbic acid (AA). It is considered that the fluorescence enhancement of the Tb3+–E–BSA–AA–SDS system originates not only from the hydrophobic microenvironment provided by BSA–SDS, but also from the energy transfer from BSA to Tb3+ in this system. Therefore, a new fluorescence method for the determination of protein concentrations as low as 1.3 × 10?9 g mL?1 BSA is established using Tb3+–epinephrine complex as probe. The method has been applied for the determination of BSA and human serum albumin in actual samples, and the results obtained are satisfactory. Compared with other fluorescence methods, this method is simpler and more sensitive for the determination of protein. The mechanism of the fluorescence enhancement of the system is studied in detail. Copyright © 2009 John Wiley & Sons, Ltd. 相似文献
18.
The function of photosystem (PS)II during desiccation and exposure to high photon flux density (PFD) was investigated via analysis of chlorophyll fluorescence in the desert resurrection plant Selaginella lepidophylla (Hook. and Grev.) Spring. Exposure of hydrated, physiologically competent stems to 2000 mol · m–2 · s–1 PFD caused significant reductions in both intrinsic fluorescence yield (FO) and photochemical efficiency of PSII (FV/FM) but recovery to pre-exposure values was rapid under low PFD. Desiccation under low PFD also affected fluorescence characteristics. Both FV/FM and photochemical fluorescence quenching remained high until about 40% relative water content and both then decreased rapidly as plants approached 0% relative water content. In contrast, the maximum fluorescence yield (FM) decreased and non-photochemical fluorescence quenching increased early during desiccation. In plants dried at high PFD, the decrease in FV/FM was accentuated and FO was reduced, however, fluorescence characteristics returned to near pre-exposure values after 24-h of rehydration and recovery at low PFD. Pretreatment of stems with dithiothreitol, an inhibitor of zeaxanthin synthesis, accelerated the decline in FV/FM and significantly increased FO relative to controls at 925 mol · m–2 · s–1 PFD, and the differences persisted over a 3-h low-PFD recovery period. Pretreatment with dithiothreitol also significantly decreased non-photochemical fluorescence quenching, increased the reduction state of QA, the primary electron acceptor of PSII, and prevented the synthesis of zeaxanthin relative to controls when stems were exposed to PFDs in excess of 250 mol · m–2 · s–1. These results indicate that a zeaxanthin-associated mechanism of photoprotection exists in this desert pteridophyte that may help to prevent photoinhibitory damage in the fully hydrated state and which may play an additional role in protecting PSII as thylakoid membranes undergo water loss.Abbreviations and Symbols DTT
dithiothreitol
- EPS
epoxidation state
- FO
yield of instantaneous fluorescence at open PSII centers
- FM
maximum yield of fluorescence at closed PSII centers induced by saturating light
- FM
FM determined during actinic illumination
- FV
yield of variable fluorescence (FM-FO)
- FV/FM
photochemical efficiency of PSII
- qP
photochemical fluorescence quenching
- qNP
non-photochemical fluorescence quenching of Schreiber et al. (1986)
- NPQ
non-photochemical fluorescence quenching from the Stern-Volmer equation
- PFD
photon flux density
- RWC
relative water content
This paper is based on research done while W.G.E. was on leave of absence at Duke University during the fall of 1990. We would like to thank Dan Yakir, John Skillman, Steve Grace, and Suchandra Balachandran and many others at Duke University for their help and input with this research. Dr. Barbara Demmig-Adams provided zeaxanthin for standard-curve purposes. 相似文献
19.
Márcio H.L Arndt Marcelo M Santoro 《Comparative biochemistry and physiology. Part B, Biochemistry & molecular biology》1998,119(4):667-675
The hemoglobin of Biomphalaria glabrata was purified to homogeneity by gel filtration column followed by anion exchange chromatography. The dissociation products were analyzed by a 5–15% gradient polyacrylamide gel electrophoresis containing sodium dodecyl sulfate (SDS-PAGE) giving a band of 270 kDa and a band of 180 kDa after reduction with β-mercaptoethanol. The same profile was obtained in a 3.5% agarose gel electrophoresis containing SDS (SDS-AGE) but showed additional bands of higher molecular weight. These bands were proposed to be monomers, dimers and trimers, since they showed a good correlation in a plot of Rf versus log Mr. After partial reduction in a two-dimensional SDS-AGE, the proposed monomers and dimers produced two and four bands, respectively, likely indicating one to four chains crosslinked by disulfide bridges. Digestion with four different proteases yielded several equivalent fragments with molecular weights multiples of its minimum molecular weight (17.7 kDa). The circular dichroism spectrum of the protein showed a characteristic high α-helix content (70%). It was proposed that this hemoglobin is a pentamer with a molecular weight of aproximately 1.8×103 kDa, assembled by five 360-kDa subunits, each formed by two 180-kDa chains linked in pairs by disulfide bridges and each of these chains, in turn, comprised by ten heme binding domains linked in tandem. These data are compared to the published information for other planorbid extracellular hemoglobins. 相似文献
20.
Henryk Szmacinski Wieslaw Wiczk Mayer N. Fishman Peggy S. Eis Joseph R. Lakowicz Michael L. Johnson 《European biophysics journal : EBJ》1996,24(3):185-193
We have examined the fluorescence intensity decays of oxytocin and [Arg8]-vasopressin resulting from the single tyrosyl residue in each peptide, and the intensity decay of the Asu 1,6-analogues in which the disulfide bridge is substituted by a CH2-CH2 bridge. Viscosity-dependent steady state and intensity decay measurements indicated that fluorescence resonance energy transfer (FRET) from tyrosyl phenol to the disulfide bridge is responsible for the decrease in fluorescence relative to the Asu-analogues. The frequency-domain phase and modulation data for the tyrosyl donor were interpreted in terms of fluorescence resonance energy transfer (FRET) to the weakly absorbing disulfide bridge and a distribution of donor-to-acceptor distances. Energy transfer efficiencies were determined from both time-resolved and steady-state measurements. Fitting the frequency-domain phase and modulation data to a Gaussian distance distribution indicated that the average inter-chromophoric distance (Rav) is similar in both compounds, Rav=7.94 Å for oxytocin and Rav = 8.00 Å for vasopressin. However, the width of the distance distribution is narrower for vasopression (hw =2.80 Å) than for oxytocin (hw =3.58 Å), which is consistent with restriction of the tyrosine phenol motion due to its stacking with the Phe3 side chain of vasopressin. Finally, the recovered distance distribution functions are compared with histograms describing the distance between the chromophores during the course of long, in vacuo, molecular dynamics runs using the computer program CHARMm and the QUANTA 3.0 parameters.Abbreviations AVP
[Arg8]-vasopressin
- FRET
fluorescence resonance energy transfer
- FD
frequency-domain
- D
donor
- A
acceptor
- DTT
dithiothreitol
Correspondence to: J. R. Lakowicz 相似文献