Spectral signatures of five hydroxymethyl chlorophyll a derivatives chemically derived from chlorophyll b or chlorophyll f

Photosynthesis Research - Chlorophylls (Chls) are pigments involved in light capture and light reactions in photosynthesis. Chl a, Chl b, Chl d, and Chl f are characterized by unique absorbance...     

Organization of chlorophyll a in bilayer membranes. The chlorophyll a/dimyristoylphosphatidylcholine system

In order to investigate the relative importance of the hydrophobic and headgroup interactions of chlorophyll a in phospholipid bilayers, we have carried out differential scanning calorimetry (DSC) and deuterium (²H) and phosphorus (³¹P) nuclear magnetic resonance (NMR) experiments on the multilamellar system of chlorophyll a in dimyristoylphosphatidylcholine (DMPC). Compared to the phytol chain of chlorophyll and the previously reported distearoylphosphatidylcholine (DSPC), the acyl chains of DMPC are shorter in length by three and four carbons, respectively. A lowering in the phase-transition temperature was observed for the DMPC multilayers in the presence of chlorophyll a in the DSC thermograms and in the ³¹P chemical shift anisotropy measurements. These results, together with data on hydrophobic interactions as measured by ²H-NMR and on headgroup interactions as evidenced from ³¹P-NMR, suggest a phase diagram for the chlorophyll a/DMPC system in which phase separation readily occurs between a chlorophyll-rich compound phase and a chlorophyll-poor phospholipid phase. Compound formation appears to be important in the stabilization of chlorophyll a in bilayers with shorter chains.     

Analysis of the chlorophyll biosynthetic system in a chlorophyll b-less barley mutant

The chlorophyll b-less barley (Hordeum vulgare L.) mutant chlorina 2807 allelic to the well-known barley mutant chlorina f2 was studied. 5-Aminolevulinic acid at saturating concentration (40 mM) was introduced into postetiolated leaves of the mutant and its wild type, and the protochlorophyllide accumulation in the dark was measured. It was found that the activity of the enzyme system transforming 5-aminolevulinic acid into protochlorophyllide was the same in both types of plants. The activity of esterifying enzymes that catalyze attachment of phytol to chlorophyllide was analyzed by infiltration of exogenous chlorophyllides a and b into etiolated leaves. The reaction was shown to have close rates in the mutant and wild-type plants. In very early stages of greening of etiolated leaves, when the apoproteins of the light-harvesting complexes are not yet formed, appearance of chlorophyll b was clearly recorded in the wild-type plants, while in the mutant chlorina 2807 no indications of chlorophyll b were detected in any stage of greening. On the other hand, in the mutant as well as in the wild type an active reverse conversion of chlorophyll b into chlorophyll a was possible. It is concluded that (a) in the mutant chlorina 2807 the ability of the biosynthetic system to transform 5-aminolevulinic acid to chlorophyll a is fully preserved, (b) in the mutant the enzymes converting chlorophyll a into chlorophyll b are most likely absent or damaged, (c) the conversion of chlorophyll a into chlorophyll b and the reverse conversion of chlorophyll b into chlorophyll a are performed by different enzymes.     

Evidence for a light-harvesting chlorophyll a-protein complex in a chlorophyll b-less barley mutant

Chloroplasts of a chlorophyll (Chl) b-less barley mutant were solubilized with digitonin and fractionated by polyacrylamide gel electrophoresis with sodium deoxycholate in the running buffer. By this procedure, in contrast to using sodium dodecylsulfate (SDS) for solubilization, a Chl a-protein analogous to the major light-harvesting Chl a-b protein complex from wildtype chloroplasts was recovered. This mutant Chl a-protein comprises about fifty percent of the total Chl a, and is very similar in carotenoid, amino acid, protein and polypeptide composition to the major wildtype antenna Chl a-b protein. The only major differences we have found is its instability in the presence of SDS and sensitivity to protease action. Even with deoxycholate, the mutant Chl a complex often dissociates during electrophoresis into two green bands. The lack of Chl b appears to affect the normal organization of Chl a and protein in such a way as to render the complex more unstable.CIW-DPB No. 917.     

Four universal forms of chlorophyll a

We have matched the red absorption band measured at −196 C in a variety of chloroplast preparations with four major component curves representing forms of chlorophyll a having peaks at 661.6, 669.6, 677.1, and 683.7 nanometers. Chloroplast fractions enriched in one or the other of the two photochemical systems both contain these four major components, but system 1 preparations contain relatively more chlorophyll a 684. Chlorophyll a 677 and chlorophyll a 684 have greater bandwidths in system 1. Bands at longer wavelengths near 693 and 704 nanometers also often occur, but with far smaller heights than the above major bands. The longer wavelength bands are more common in system 1 than in system 2. In system 1 the half-widths of the four major bands in typical spectra average 11.3, 10.0, 10.3, and 10.8 nanometers while in system 2 they are 11.6, 9.8, 9.4, and 9.6 nanometers. Some spectra with sharper and some with wider bands were found, but the wavelengths were identical.     

Identification of the 7-hydroxymethyl chlorophyll a reductase of the chlorophyll cycle in Arabidopsis

The interconversion of chlorophyll a and chlorophyll b, referred to as the chlorophyll cycle, plays a crucial role in the processes of greening, acclimation to light intensity, and senescence. The chlorophyll cycle consists of three reactions: the conversions of chlorophyll a to chlorophyll b by chlorophyllide a oxygenase, chlorophyll b to 7-hydroxymethyl chlorophyll a by chlorophyll b reductase, and 7-hydroxymethyl chlorophyll a to chlorophyll a by 7-hydroxymethyl chlorophyll a reductase. We identified 7-hydroxymethyl chlorophyll a reductase, which is the last remaining unidentified enzyme of the chlorophyll cycle, from Arabidopsis thaliana by genetic and biochemical methods. Recombinant 7-hydroxymethyl chlorophyll a reductase converted 7-hydroxymethyl chlorophyll a to chlorophyll a using ferredoxin. Both sequence and biochemical analyses showed that 7-hydroxymethyl chlorophyll a reductase contains flavin adenine dinucleotide and an iron-sulfur center. In addition, a phylogenetic analysis elucidated the evolution of 7-hydroxymethyl chlorophyll a reductase from divinyl chlorophyllide vinyl reductase. A mutant lacking 7-hydroxymethyl chlorophyll a reductase was found to accumulate 7-hydroxymethyl chlorophyll a and pheophorbide a. Furthermore, this accumulation of pheophorbide a in the mutant was rescued by the inactivation of the chlorophyll b reductase gene. The downregulation of pheophorbide a oxygenase activity is discussed in relation to 7-hydroxymethyl chlorophyll a accumulation.     

Use of the atLEAF+ chlorophyll meter for a nondestructive estimate of chlorophyll content

At present, chlorophyll meters are widely used for a quick and nondestructive estimate of chlorophyll (Chl) contents in plant leaves. Chl meters allow to estimate the Chl content in relative units - the Chl index (CI). However, using such meters, one can face a problem of converting CI into absolute values of the pigment content and comparing data acquired with different devices and for different plant species. Many Chl meters (SPAD-502, CL-01, CCM-200) demonstrated a high degree of correlation between the CI and the absolute pigment content. A number of formulas have been deduced for different plant species to convert the CI into the absolute value of the photosynthetic pigment content. However, such data have not been yet acquired for the atLEAF+ Chl meter. The purpose of the present study was to assess the applicability of the atLEAF+ Chl meter for estimating the Chl content. A significant species-specific exponential relationships between the atLEAF value (corresponding to CI) and extractable Chl a, Chl b, Chl (a+b) for Calamus dioicus and Cleistanthus sp. were shown. The correlations between the atLEAF values and the content of Chl a, Chl b, and Chl (a+b) per unit of leaf area was stronger than that per unit of dry leaf mass. The atLEAF value- Chl b correlation was weaker than that of atLEAF value-Chl a and atLEAF value-Chl (a+b) correlations. The influence of light conditions (Chl a/b ratio) on the atLEAF value has been also shown. The obtained results indicated that the atLEAF+ Chl meter is a cheap and convenient tool for a quick nondestructive estimate of the Chl content, if properly calibrated, and can be used for this purpose along with other Chl meters.     

The photosynthetic response to a shift in the chlorophyll a to chlorophyll B ratio of chlorella

The chlorophyll a:b ratio was shifted in Chlorella vannielii by varying the illuminance under which the cells were cultured—the ratio increased from 2.9, 3.0, 4.0, and 4.8 to 6.2, respectively, at 100, 300, 900, 2,700 and 6,000 foot candles. The 6,000-foot candle cells retained an optimal growth rate at the chlorophyll a:b ratio of 6.2 which was the upper limit of normal growth. Comparisons were made between the 300-and 6,000-foot candle cultures to determine the significance to the photosynthetic mechanism of a shift in the chlorophyll a:b ratio.     

Dichroism and polarized fluorescence of chlorophyll a, chlorophyll c and bacteriochlorophyll a dissolved in liquid crystals

Three photosynthetic pigments were studied: chlorophyll a, chlorophyll c and bacteriochlorcphyll a in nematic liquid crystal matrixes. The polarized absorption and fluorescence spectra as a function of the electric field have been measured. From the polarized components of the absorption A( parallel) and A( perpendicular) of the pigments in liquid crystals two reduced components A(x) and A(y) are calculated (x and y are the direction of the axis which is going through the second, fourth pyrrol rings, and the first, third rings, respectively). From these results the orientation of chlorophylls in liquid crystals and the configuration of the transition moments in the skeleton of the pigment molecules were determined.     

In vitro synthesis of chlorophyll a in the dark triggers accumulation of chlorophyll a apoproteins in barley etioplasts

An in vitro translation system using lysed etioplasts was developed to test if the accumulation of plastid-encoded chlorophyll a apoproteins is dependent on the de novo synthesis of chlorophyll a. The P700 apoproteins, CP47 and CP43, were not radiolabeled in pulsechase translation assays employing lysed etioplasts in the absence of added chlorophyll precursors. When chlorophyllide a plus phytylpyrophosphate were added to lysed etioplast translation assays in the dark, chlorophyll a was synthesized and radiolabeled P700 apoproteins, CP47 and CP43, and a protein which comigrates with D1 accumulated. Chlorophyllide a or phytylpyrophosphate added separately to the translation assay in darkness did not induce chlorophyll a formation or chlorophyll a apoprotein accumulation. Chlorophyll a formation and chlorophyll a apoprotein accumulation were also induced in the lysed etioplast translation system by the photoreduction of protochlorophyllide to chlorophyllide a in the presence of exogenous phytylpyrophosphate. Accumulation of radiolabeled CP47 was detectable when very low levels of chlorophyll a were synthesized de novo (less than 0.01 nmol/10(7) plastids), and radiolabel increased linearly with increasing de novo chlorophyll a formation. Higher levels of de novo synthesized chlorophyll a were required prior to detection of radiolabel incorporation into the P700 apoproteins and CP43 (greater than 0.01 nmol/10(7) plastids). Radiolabel incorporation into the P700 apoproteins, CP47 and CP43, saturated at a chlorophyll a concentration which corresponds to 50% of the etioplast protochlorophyllide content (0.06 nmol of chlorophyll a/10(7) plastids).     

脱镁叶绿酸a单加氧酶与叶绿素降解

近年来叶绿素降解的分子调控机制颇受关注,脱镁叶绿酸a单加氧酶（PaO）作为催化叶绿素卟啉环开环成为四元线性吡咯衍生物的关键酶,成为研究的焦点。本文从PaO的结构与生理功能出发,着重介绍了近期在PaO表达调控方面的最新研究进展,包括PaO突变体中叶绿素降解的反馈机制;去磷酸化作用与PaO的活性调节;非黄化突变体AtNYE1基因、蛋白激酶基因rlpk2、持绿突变体基因sgr的表达与PaO活性之间的关系,并且简要归纳了PaO研究的发展过程与趋势、存在的问题和争议。     

Determination of relative chlorophyll binding affinities in the major light-harvesting chlorophyll a/b complex

The major light-harvesting complex (LHCIIb) of photosystem II can be reconstituted in vitro from its recombinant apoprotein in the presence of a mixture of carotenoids and chlorophylls a and b. By varying the chlorophyll a/b ratio in the reconstitution mixture, the relative amounts of chlorophyll a and chlorophyll b bound to LHCIIb can be changed. We have analyzed the chlorophyll stoichiometry in recombinant wild type and mutant LHCIIb reconstituted at different chlorophyll a/b ratios in order to assess relative affinities of the chlorophyll-binding sites. This approach reveals five sites that exclusively bind chlorophyll b. Another site exhibits a slight preference of chlorophyll b over chlorophyll a. The remaining six sites are filled preferentially with chlorophyll a but also tolerate chlorophyll b when this is offered at a large excess. Three of these chlorophyll a-affine sites could be assigned to distinct positions defined by the three-dimensional LHCIIb structure. Exclusive chlorophyll b sites complemented by chlorophyll a sites that are selective only to a certain extent are consistent with the observation that chlorophyll b but not chlorophyll a is essential for reconstituting stable LHCIIb. These data offer an explanation why a rather constant chlorophyll a/b ratio is observed in native LHCIIb despite the apparent promiscuity of some binding sites.