Advanced plant physiology. Photomorphogenesis Light mediated photoresponse processes. *...
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Advanced plant physiology
Photomorphogenesis Light mediated photoresponse processes. * Photosynthesis --- a biochemical process * Photoperiodic control of reproductive behavior
* Nonphotosynthetic photoreceptor ---- phytochrome ---- Protochlorophyll, the synthesis of chlorophyll and development of photosynthetic apparatus.
Phytochrome---as photoreceptor, known for long time. lettuce seed germination of certain variety are sensitive to light, (the grand rapid variety) Did absorbance test of spectro,
Phytochorme --- by giving different wavelength to get 50% germination. showed that main region of germination is in the red region.
1946 Bonner etc. studied the light interruption by using photographic projector. 1952 Hendrick etc reinvestigate lettuce seed. --found low germination in dark. give red light --- increase germination. red, followed by far-red light inhibit germination. Red -- Far red --- red again --- increase again.
a similar effect on Xanthium, a short day plant.
Long night ---- flowering
red light interrupt --- no flowering Red light interrupt long night followed by far red -- flowering.
1952-1959 try to get some physiological evidence. ---- Butler, norris, Siegleman & Hendrick Irradiate with red light ---- decrease in absorbance at 660 nm---- increase in absorbance at 730 nm Irradiate with far red light---- increase in absorbance at 660 nm---- decrease in absorbance at 730 nm
The compound is sensitive to heat, must be protein in nature, same conclusion as Red, White Pr Pfr Far red, DarkStructure: Protein + Chromophore, similar to phycobilin in algae.
Effect of light: Act on chromophore first then act on protein. idea: the sequence of event suggest that possibly changes of the conformation (configuration) between cis and trans form of the phytochrome molecules.
Start with 100% Pr then irradiate with red light at 10 sec. interval it begin to convert Pr into Pfr at a first order reaction rate:
at some point, the absorption are due to both Pr and Pfr. ex. 80% convert to pfr, 20% to pr -- it is calledPhotostationary state after that, give more light, still no more effect.
at 698 nm wavelength, Pr=50% and Pfr=50% if, Pfr were irradiated with far-red, it reverse to 98% Pr and 1-2% Pfr a dose response to red or far-red or attainment of photostationary state can be shown. see figure.
OD 730OD 660730 nmOD660 nmOD
13312&4RedSat1 2. 3698nm23. FRSat3 4. -369846607302&4
What are various light do on phytochromesun light = --- Pfr relatively high fluorescent ---- high Pfr (tube light)Incandescent --- low Pfr (more heat)Dark reversion: Pfr convert to Pr in dark
to many plants, when red irradiated treatment Pr convert to Pfr. then Pfr start to decay or denatured, and no more dark reversion.ex. in corn seedlings, only 20% dark reversion from Pfr, it is a common phenomeon in monocot.
Phytochrome. method of detection --- in vivo, through tissue segment. Sample harvest in dim green light. Sample harvest in green light as safe light, or in total darkness.
The change from Pr to Pfr transformation reflect changes in protein conformation possibly that a single photoreceptor pigment could exist in two photoconvertible forms:
Pr have an action maximun near 660 nm, and Pfr near 730 nm. absorption of light by either form converted it to the other form.
The pigment acting as a photoreceptor and that its absorption maximum must be close to the action maximum. Predict that the pigment would be blue in colour.
The low energy requirement suggested that the photoaction brings about a change in molecular configuration rather athan a transfer of energy to another system.
* Difficulty of detecting phytochrome.1. Low concentration of the pigment in plant tissue with very high molar extinction coefficients.
A= -logT= abcBeers Law : ln I/Io = -KbcIo= Intensity of incidence radiationI= Change in intensity of IoT= Transmittance = I/Ioa= extinction coefficient or = absorbancy index = K / 2.303 to give base10 log (log10 ()) =cm3mg-1cm-1 or cm2mg-1b= optical path = 1cmA is proportional to c
E= molar extinction coefficient = M-1cm-1 or liter mole-1 cm-1 E: A. 1mg/ml solution of BSA in neutral phosphate buffer has an A280 of 0.6 If the molecular weight of BAS is 66,000 what is the molar extinction coefficient? Beers Law concept: A is proportional to c: If light is absorbed by a substance in solution, a (constant) is proportional to the concentration of the substance in solution.
* Difficulty of detecting phytochrome.2. The overriding absorbance of chlorophyll has obscured the measurement of phytochrome in green tissue.
Isolation and purification of phytochrome.methods for extracting proteins were used in the isolation. dark grown seedlings.Phytochrome, a protein with a chromophoric (pigment) group, low radiant energy needed to bring about.
a physiological response suggested a need for amplication analogous to the functioning of an enzyme.trace amount of substance are usually physiologically active by virtue of binding to a protein.
attempt to isolate phytochrome as a protein were successful, but subsequent purification require considerable time. isolate phytochrome chromophere directly from plant tissue by method used in pigment chemistry have not yet been successful.
Purification of phytochrome has been satisfactorily achieved to date only from dark grown monocot. seedlings (avena sativa, oat). pH is a key factor for the purification. above pH 7.3 in supernatant contained the maximal extractable pigment. at pH 6.2 or below, only in sediment.
In supernatantpH 7.3In sedimentpH 6.2
The intracellular location of phytochrome has not been systematically pursued.
New satisfactory procedure for purification is available, mostly monocot, dark grown seedling, spinach, Mesotaenium (green algae) Sphaerocarpus (a liverwort).
Absorption of light by the chromophroe brings about a change in conformation of the protein, which passes through several intermediate to the final form.
Denaturation .... Photoreversibility of phytochrome are lost on denaturation is an indicative of a dependence on interaction of the
chromophore with the native protein. Pfr form is more susceptible to urea, proteolytic enzymes and -SH reagent than Pr.
Higher plant is a captive of its environment and must be provided with sensor to appraise variables of its milieu.
light is one of the most important variables. Many aspects of the growth and development of higher plants are under control of phytochrome. ex. flowering, other aspects of phytochrome mediated process are:
1. Etiolation --- an assemblage of several responses, including control of phyocotyl length, leaf expansion and plumular hook opening. red light induced a reduction in hypocotyl length., a growth inhibition and expansion of leaves --- a growth promotion.
2. Internode length control --- mediated by phytochrome. These results are similar to those for reversal of etiolation. The location of the photoreceptor for the regulation of nternode length was in the internode itself.
3. Biochemical reaction under phytochrome control are.
a. coloration of the tomato cuticle is a biochemical synthesis under phytochrome control resulting from the deposition of a yet unknown pigment into the cuticle of the fruit.
b. Flavonoid synthesis in many plants requires light and is under phytochrome control. Pfr enhance the synthesis.Flavonoid
c. Anthocyanin in apple skin is a case of phytochrome control. apple skin forms anthocyanin when exposed to light and accumulates aldehyde and alcohol when held in darkness.precursorlightdarkanthocyaninaldehyde & alcohol
c. substrate for anthocyanin synthesis in light, if not utlized, forms aldehyde and alcohol. pigment is banana pseudo-stem possible phytochrome mdeiated.
d. development and differentiation of plastids of etiolated leaves of higher plants is light dependent.
protochlorophyll --- chlorophyll
prolamella bodies --- lamella & grana formation.
e. NADP-dependent triose- phosphate dehydrogenase and chloroplast protein. (Marcus, Margulis). Prolognged far-red irradiation == brief red light effect.
f. One aspect of the phytochrome mediated effects of red light at the cellular level is the induction of plastid enzyme synthesis. By determining the magnitude of increase induced by red light