The Extra Loop Distinguishing POR from the Structurally Related Short-chain Alcohol Dehydrogenases Is Dispensable for Pigment Binding but Needed for the Assembly of Light-harvesting POR-Protochlorophyllide Complex

Reinbothe, Christiane; Lepinat, Anja; Deckers, Markus; Beck, Erwin; Reinbothe, Steffen

doi:10.1074/jbc.m209739200

Cited by 16 publications

(14 citation statements)

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“…19). Similar spectral pigment species and interconversions have been described for reconstituted PORA:PORB-lipid supracomplexes (15)(16)(17)(18). All information available at the moment suggests that the native and reconstituted complexes are structurally and functionally identical.…”

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confidence: 68%

“…Cys-1 is facing the GASSGK/LG motif, whereas Cys-3 is juxtapositioned to the YxxxK motif participating in the catalytic mechanism. Cys-2 has no counterpart in POR of pea and was therefore disregarded as Pchlide binding site, whereas Cys-4 is present between ␤-strand ␤6 and helix ␣-G and just opposite to the extra loop, establishing an oligomerization domain between the PORA and PORB (18).…”

Section: Resultsmentioning

confidence: 99%

“…PORA and PORB constitute larger light-harvesting POR:Pchlide complexes dubbed LHPP in the prolamellar body of etioplasts (15)(16)(17)(18). These complexes additionally interact with galacto-and sulfolipids and establish two spectral forms of Pchlide: Pchlide-650/ 657 and Pchlide-628/632 (the first number indicates the absorption maximum, the second shows the respective fluorescence emission maximum).…”

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confidence: 99%

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Photoprotective role of NADPH:protochlorophyllide oxidoreductase A

Buhr

Bakkouri

Valdez

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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Photoprotective role of NADPH:protochlorophyllide oxidoreductase A

Buhr

Bakkouri

Valdez

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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“…5 and 8) and previous work [25][26][27]62]. In addition to PORA and PORB and their cognate pigment substrates, the resolved supra-complex contains galacto-and sulfolipids that shift the absorption máximum of the reconstituted complex from 630 to 650 nm [25][26][27].…”

Section: Size Modula Tion Oflhppmentioning

confidence: 99%

LHPP, the light-harvesting NADPH:protochlorophyllide (Pchlide) oxidoreductase:Pchlide complex of etiolated plants, is developmentally expressed across the barley leaf gradient

et al. 2004

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NADPH:protochlorophyllide oxidoreductase is a key enzyme for the light-induced greening of etiolated angiosperm plants. In barley, two POR proteins exist termed PORA and PORB that have previously been proposed to stmcturally and functionally cooperate in terms of a higher molecular mass light-harvesting complex named LHPP, in the prolamellar body of etioplasts [Nature 397 (1999) 80]. In this study we examined the expression pattern of LHPP during seedling etiolation and de-etiolation under different experimental conditions. Our results show that LHPP is developmentally expressed across the barley leaf gradient. We further provide evidence that LHPP operates both in plants that etiolate completely before being exposed to white light and in plants that etiolate only partially and begin light-harvesting as soon as traces of light become available in the uppermost parts of the soil. As a result of light absorption, in either case LHPP converts Pchlide a to chlorophyllide (Chlide) a and in turn disintegrates. The released Chlide a, as well as Chlide b produced upon LHPP's light-dependent dissociation, which leads to the activation of the PORA as a Pchlide £-reducing enzyme, then bind to homologs of water-soluble chlorophyll proteins of Brassicaceae. We propose that these proteins transfer Chlide a and Chlide b\o the thylakoids, where their esterification with phytol and assembly into the photosynthetic membrane complexes ultimately takes place. Presumably due to the tight coupling of LHPP synthesis and degradation, as well as WSCP formation and photosynthetic membrane assembly, eflficient photo-protection is conferred onto the plant.

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“…Therefore the proposed occurrence of an LHPP in etiolated plant tissues is untenable, and its ensuing consequences and implications (9,10) for the greening process are irrelevant.…”

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Chloroplast Biogenesis 88

Kolossov

Rebeiz

2003

Journal of Biological Chemistry

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It has recently been reported that protochlorophyllide (Pchlide) b is an abundant pigment in barley etioplasts but is rather unstable, as it is rapidly converted to Pchlide a by 7-formyl reductase during pigment extraction with conventional 80% acetone (Reinbothe, S., Pollmann, S., and Reinbothe, C. The occurrence of monovinyl (MV) 1 Pchlide b in green plants was reported in 1991, and its chemical structure was ascertained by 1 H NMR and fast atom bombardment mass spectroscopy as well as by fluorescence spectroscopy (1). Later on, spectrofluorometric techniques were developed for the quantitative determination of MV-Pchlide b under various growth conditions (2). Its concentration in green cucumber cotyledons ranged from 400 to 800 pmol/100 mg of tissue protein. It was not detectable in etiolated tissues but was proposed as a logical precursor of chlorophyll b via two distinct biosynthetic routes in greening and green plant tissues (3).In 1999, Reinbothe et al. (4) proposed that in barley etioplasts, Pchlide a and Pchlide b formed a supramolecular complex with NADPH and two closely related NADPH:Pchlide oxidoreductases (POR), namely PORA and PORB. The complex was referred to as a light-harvesting POR-Pchlide complex (LHPP). It has also been suggested that LHPP contained five non-photoactive Pchlide b and one photoactive Pchlide a. Reinbothe et al. (4) conjectured that the formation of LHPP is essential for the development of the photosynthetic apparatus and conferred photoprotection upon the greening system.The above claims were rapidly refuted by two separate research groups that could not find any convincing evidence supporting the occurrence of Pchlide b in etiolated barley (5, 6). In response to the aforementioned criticism, Reinbothe et al. (7) recently refined their claims. Although they reasserted their belief in the ubiquitous occurrence of LHPP in etiolated tissues, they proposed that Pchlide b is metabolically unstable, and during conventional pigment extraction in organic solvents, it is rapidly converted to MV-Pchlide a by 7-formyl reductase, an enzyme that has been shown to be active in the conversion of chlorophyll (Chl) b to Chl a (8).In this work we evaluate the assertions of Reinbothe et al. (7) and present experimental evidence that rule out (a) the occurrence of Pchlide b in etiolated tissues and (b) its putative conversion to Pchlide a during conventional pigment extraction with 80% acetone. Therefore the proposed occurrence of an LHPP in etiolated plant tissues is untenable, and its ensuing consequences and implications (9, 10) for the greening process are irrelevant. MATERIALS AND METHODSPlant Material-Barley (Hordeum Vulgare, Hi Barley Brand) seeds were purchased from Illini FS Inc. (Urbana, IL). Germination was carried out in plastic trays containing wet vermiculate either in darkness or in a growth chamber illuminated with 1000-watt metal halide lamps (211 Watt m Ϫ2 ) under a 14-h light/10-h dark photoperiod. At midday, etiolated or green leaves were harvested after 6 -7 days of growt...

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The Extra Loop Distinguishing POR from the Structurally Related Short-chain Alcohol Dehydrogenases Is Dispensable for Pigment Binding but Needed for the Assembly of Light-harvesting POR-Protochlorophyllide Complex

Cited by 16 publications

References 46 publications

Photoprotective role of NADPH:protochlorophyllide oxidoreductase A

Photoprotective role of NADPH:protochlorophyllide oxidoreductase A

LHPP, the light-harvesting NADPH:protochlorophyllide (Pchlide) oxidoreductase:Pchlide complex of etiolated plants, is developmentally expressed across the barley leaf gradient

Chloroplast Biogenesis 88

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