The Light Dependent Reactions of Photosynthesis

Glycation of low-density lipoprotein results in the time-dependent accumulation of cholesteryl esters and apolipoprotein B-100 protein in primary human monocyte-derived macrophages Bronwyn E. Brown 1 , Imran Rashid 1 , David M. van Reyk 2 and Michael J. Davies 1,3 1 Free Radical Group, The Heart Research Institute, Camperdown, Sydney, NSW, Australia 2 Department of Health Sciences, University of Technology Sydney, NSW, Australia 3 Faculty of Medicine, University of Sydney, NSW, Australia Complications associated with diabetes are the major cause of mortality and morbidity in people with this disease. These include microvascular complications that induce damage to the retina, nephrons and peripheral nerves, and macrovascular disease that is associated with accelerated atherosclerosis (deposition of lipids in Keywords aldehydes; atherosclerosis; foam cells; human monocyte-derived macrophages; low-density lipoproteins Correspondence M. J. Davies, 114 Pyrmont Bridge Road, Camperdown, Sydney, NSW 2050, Australia Fax: +61 2 95655584 Tel: +61 2 82088900 E-mail: daviesm@hri.org.au (Received 12 December 2006, accepted 15 January 2007) doi:10.1111/j.1742-4658.2007.05699.x Nonenzymatic covalent binding (glycation) of reactive aldehydes (from glu- cose or metabolic processes) to low-density lipoproteins has been previ- ously shown to result in lipid accumulation in a murine macrophage cell line. The formation of such lipid-laden cells is a hallmark of atheroscler- osis. In this study, we characterize lipid accumulation in primary human monocyte-derived macrophages, which are cells of immediate relevance to human atherosclerosis, on exposure to low-density lipoprotein glycated using methylglyoxal or glycolaldehyde. The time course of cellular uptake of low-density lipoprotein-derived lipids and protein has been character- ized, together with the subsequent turnover of the modified apolipoprotein B-100 (apoB) protein. Cholesterol and cholesteryl ester accumulation occurs within 24 h of exposure to glycated low-density lipoprotein, and increases in a time-dependent manner. Higher cellular cholesteryl ester lev- els were detected with glycolaldehyde-modified low-density lipoprotein than with methylglyoxal-modified low-density lipoprotein. Uptake was signifi- cantly decreased by fucoidin (an inhibitor of scavenger receptor SR-A) and a mAb to CD36. Human monocyte-derived macrophages endocytosed and degraded significantly more 125 I-labeled apoB from glycolaldehyde-modified than from methylglyoxal-modified, or control, low-density lipoprotein. Dif- ferences in the endocytic and degradation rates resulted in net intracellular accumulation of modified apoB from glycolaldehyde-modified low-density lipoprotein. Accumulation of lipid therefore parallels increased endocytosis and, to a lesser extent, degradation of apoB in human macrophages exposed to glycolaldehyde-modified low-density lipoprotein. This accumula- tion of cholesteryl esters and modified protein from glycated low-density lipoprotein may contribute to cellular dysfunction and the increased atherosclerosis observed in people with diabetes, and other pathologies linked to exposure to reactive carbonyls. Abbreviations AGE, advanced glycation end-products; apoB, apolipoprotein B-100; HBSS, Hank’s balanced salt solution; HMDM, human monocyte-derived macrophage; HSA, human serum The Light-Dependent Reactions of Photosynthesis The Light-Dependent Reactions of Photosynthesis Bởi: OpenStaxCollege How can light be used to make food? When a person turns on a lamp, electrical energy becomes light energy Like all other forms of kinetic energy, light can travel, change form, and be harnessed to work In the case of photosynthesis, light energy is converted into chemical energy, which photoautotrophs use to build carbohydrate molecules ([link]) However, autotrophs only use a few specific components of sunlight Photoautotrophs can capture light energy from the sun, converting it into the chemical energy used to build food molecules (credit: Gerry Atwell) What Is Light Energy? The sun emits an enormous amount of electromagnetic radiation (solar energy) Humans can see only a fraction of this energy, which portion is therefore referred to as “visible 1/12 The Light-Dependent Reactions of Photosynthesis light.” The manner in which solar energy travels is described as waves Scientists can determine the amount of energy of a wave by measuring its wavelength, the distance between consecutive points of a wave A single wave is measured from two consecutive points, such as from crest to crest or from trough to trough ([link]) The wavelength of a single wave is the distance between two consecutive points of similar position (two crests or two troughs) along the wave Visible light constitutes only one of many types of electromagnetic radiation emitted from the sun and other stars Scientists differentiate the various types of radiant energy from the sun within the electromagnetic spectrum The electromagnetic spectrum is the range of all possible frequencies of radiation ([link]) The difference between wavelengths relates to the amount of energy carried by them 2/12 The Light-Dependent Reactions of Photosynthesis The sun emits energy in the form of electromagnetic radiation This radiation exists at different wavelengths, each of which has its own characteristic energy All electromagnetic radiation, including visible light, is characterized by its wavelength Each type of electromagnetic radiation travels at a particular wavelength The longer the wavelength (or the more stretched out it appears in the diagram), the less energy is carried Short, tight waves carry the most energy This may seem illogical, but think of it in terms of a piece of moving a heavy rope It takes little effort by a person to move a rope in long, wide waves To make a rope move in short, tight waves, a person would need to apply significantly more energy The electromagnetic spectrum ([link]) shows several types of electromagnetic radiation originating from the sun, including X-rays and ultraviolet (UV) rays The higher-energy waves can penetrate tissues and damage cells and DNA, explaining why both X-rays and UV rays can be harmful to living organisms Absorption of Light Light energy initiates the process of photosynthesis when pigments absorb the light Organic pigments, whether in the human retina or the chloroplast thylakoid, have a narrow range of energy levels that they can absorb Energy levels lower than those represented by red light are insufficient to raise an orbital electron to a populatable, excited (quantum) state Energy levels higher than those in blue light will physically tear the molecules apart, called bleaching So retinal pigments can only “see” (absorb) 700 nm to 400 nm light, which is therefore called visible light For the same reasons, plants pigment molecules absorb only light in the wavelength range of 700 nm to 400 nm; plant physiologists refer to this range for plants as photosynthetically active radiation The visible light seen by humans as white light actually exists in a rainbow of colors Certain objects, such as a prism or a drop of water, disperse white light to reveal the colors to the human eye The visible light portion of the electromagnetic spectrum shows the rainbow of colors, with violet and blue having shorter wavelengths, and therefore higher energy At the other end of the spectrum toward red, the wavelengths are longer and have lower energy ([link]) 3/12 The Light-Dependent Reactions of Photosynthesis The colors of visible light not carry the same amount of energy Violet has the shortest wavelength and therefore carries the most energy, whereas red has the longest wavelength and carries the least amount of energy (credit: modification of work by NASA) Understanding Pigments Different kinds of pigments exist, and each has evolved to absorb only certain wavelengths (colors) of visible light Pigments reflect or transmit the wavelengths they cannot absorb, making them appear in the corresponding color Chlorophylls and carotenoids are the two major classes of photosynthetic pigments found in plants and algae; each class has multiple types of pigment molecules There are five major chlorophylls: a, b, c and d and a related molecule found in prokaryotes called bacteriochlorophyll Chlorophyll a and ...Enzymes for the NADPH-dependent reduction of dihydroxyacetone and D-glyceraldehyde and L-glyceraldehyde in the mould Hypocrea jecorina Janis Liepins 1,2 , Satu Kuorelahti 1 , Merja Penttila ¨ 1 and Peter Richard 1 1 VTT Biotechnology, Espoo, Finland 2 University of Latvia, Institute of Microbiology and Biotechnology, Riga, Latvia Dihydroxyacetone (DHA), d-glyceraldehyde and l-glyceraldehyde can be reduced using NADPH as a cofactor to form glycerol and NADP. Enzymes cataly- sing this reaction are generally called NADP:glycerol dehydrogenases. NADP:glycerol dehydrogenase activ- ity is common in moulds and filamentous fungi. Enzymes from different species of filamentous fungi have been purified and characterized. The enzymes purified from Aspergillus niger [1] and Aspergillus nidu- lans [2] catalyse the reversible reaction from glycerol and NADP to DHA and NADPH. For the A. niger enzyme, an equilibrium constant of 3.1–4.6 · 10 )12 m was estimated for the reaction: Glycerol þ NADP Ð DHA þ NADPH þ H þ A glycerol dehydrogenase with slightly different prop- erties was described in Neurospora crassa, where d-glyceraldehyde was the preferred substrate over DHA in the reductive reaction. This enzyme was also reversible, i.e. it showed activity with glycerol and NADP [3]. The purified glycerol dehydrogenases from A. nidulans and A. niger also showed low activity with d-glyceraldehyde; however, DHA was the preferred substrate [2]. The A. niger enzyme was commercially available as a partly purified preparation, and partial amino acid sequence s were available [4]. Keywords dihydroxyacetone; glycerol dehydrogenase; Hypocrea jecorina; L-glyceraldehyde; NADP- specific glycerol dehydrogenase Correspondence P. Richard, VTT, Tietotie 2, Espoo, PO Box 1000, 02044 VTT, Finland Fax: +358 20 722 7071 Tel: +358 20 722 7190 E-mail: Peter.Richard@vtt.fi (Received 4 May 2006, revised 7 July 2006, accepted 17 July 2006) doi:10.1111/j.1742-4658.2006.05423.x The mould Hypocrea jecorina (Trichoderma reesei) has two genes coding for enzymes with high similarity to the NADP-dependent glycerol dehy- drogenase. These genes, called gld1 and gld2, were cloned and expressed in a heterologous host. The encoded proteins were purified and their kinetic properties characterized. GLD1 catalyses the conversion of d-glyceralde- hyde and l-glyceraldehyde to glycerol, whereas GLD2 catalyses the con- version of dihydroxyacetone to glycerol. Both enzymes are specific for NADPH as a cofactor. The properties of GLD2 are similar to those of the previously described NADP-dependent glycerol-2-dehydrogenases (EC 1.1.1.156) purified from different mould species. It is a reversible enzyme active with dihydroxyacetone or glycerol as substrates. GLD1 resembles EC 1.1.1.72. It is also specific for NADPH as a cofactor but has otherwise completely different properties. GLD1 reduces d-glyceraldehyde and l-glyceraldehyde with similar affinities for the two substrates and sim- ilar maximal rates. The activity in the oxidizing reaction with glycerol as substrate was under our detection limit. Although the role of GLD2 is to facilitate glycerol formation under osmotic stress conditions, we hypothes- ize that GLD1 is active in pathways for sugar acid catabolism such as d-galacturonate catabolism. Abbreviations DHA, dihydroxyacetone; DHAP, dihydroxyacetone phosphate. FEBS Journal 273 (2006) 4229–4235 ª 2006 The Authors Journal compilation ª 2006 FEBS 4229 Glycerol dehydrogenases have different functions in Light-induced reactions of Escherichia coli DNA photolyase monitored by Fourier transform infrared spectroscopy Erik Schleicher 1, *, Benedikt Heßling 2 , Viktoria Illarionova 1 , Adelbert Bacher 1 , Stefan Weber 3 , Gerald Richter 1, † and Klaus Gerwert 2 1 Lehrstuhl fu ¨ r Organische Chemie und Biochemie, Technische Universita ¨ tMu ¨ nchen, Germany 2 Lehrstuhl fu ¨ r Biophysik, Ruhr-Universita ¨ t-Bochum, Germany 3 Freie Universita ¨ t Berlin, Fachbereich Physik, Berlin, Germany Cyclobutane pyrimidine dimers (Pyr<>Pyr) and pyri- midine–pyrimidone (6–4) photoproducts are the predo- minant structural modifications resulting from exposure of DNA to ultraviolet light [1,2]. The structure of Pyr<>Pyr was elucidated by Blackburn and Davies already 40 years ago [3,4]. Both photoproducts result from 2p+2p cyclo-additions. The potentially mutagenic or lethal modifications [5] must be repaired in order to ensure cell survival and genetic stability. This can be effected by excision-repair or by photoreactivation Keywords DNA photolyase; DNA repair; FT-IR; pyrimidine dimer; stable-isotope labelling Correspondence G. Richter, School of Biological and Chemical Sciences, University of Exeter, Stocker Rd, Exeter, EX4 4QD, UK Fax: +44 1392 26 3434 Tel: +44 1392 26 3494 E-mail: g.richter@exeter.ac.uk K. Gerwert, Lehrstuhl fu ¨ r Biophysik, Ruhr-Universita ¨ t-Bochum, Universita ¨ tsstr. 150, 44780 Bochum, Germany Fax: +49 2343 21 4238 Tel: +49 2343 22 4461 E-mail: gerwert@bph.ruhr-uni-bochum *Present address Freie Universita ¨ t Berlin, Fachbereich Physik, Arnimallee 14, 14195 Berlin, Germany †Present address School of Biological and Chemical Sciences, University of Exeter, UK (Received 9 December 2004, revised 10 February 2005, accepted 16 February 2005) doi:10.1111/j.1742-4658.2005.04617.x Cyclobutane-type pyrimidine dimers generated by ultraviolet irradiation of DNA can be cleaved by DNA photolyase. The enzyme-catalysed reaction is believed to be initiated by the light-induced transfer of an electron from the anionic FADH ) chromophore of the enzyme to the pyrimidine dimer. In this contribution, first infrared experiments using a novel E109A mutant of Escherichia coli DNA photolyase, which is catalytically active but unable to bind the second cofactor methenyltetrahydrofolate, are described. A stable blue-coloured form of the enzyme carrying a neutral FADH radical cofactor can be interpreted as an intermediate analogue of the light-driven DNA repair reaction and can be reduced to the enzymatically active FADH ) form by red-light irradiation. Difference Fourier transform infra- red (FT-IR) spectroscopy was used to monitor vibronic bands of the blue radical form and of the fully reduced FADH ) form of the enzyme. Preliminary band assignments are based on experiments with 15 N-labelled enzyme and on experiments with D 2 O as solvent. Difference FT-IR mea- surements were also used to observe the formation of thymidine dimers by ultraviolet irradiation and their repair by light-driven photolyase catalysis. This study provides the basis for future time-resolved FT-IR studies which are aimed at an elucidation of a detailed molecular picture of the light- driven DNA repair process. Abbreviations DTT, dithiothreitol; FT-IR, Fourier transform infrared; MTHF, 5,10-methenyltetrahydrofolylpolyglutamate; Pyr<>Pyr, cyclobutane pyrimidine dimmer. FEBS Journal 272 (2005) 1855–1866 ª 2005 FEBS 1855 mediated by DNA photolyases. Specifically, photolyases catalyse the light-driven cleavage of the cyclobutane ring of tricyclic pyrimidine dimers, and (6–4) photolyases cleave the pyrimidine (6–4) pyrimidone Analysis of the CK2-dependent phosphorylation of serine 13 in Cdc37 using a phospho-specific antibody and phospho-affinity gel electrophoresis Yoshihiko Miyata and Eisuke Nishida Department of Cell and Developmental Biology, Graduate School of Biostudies, Kyoto University, Japan Protein kinases play pivotal roles in cellular signal transduction systems. Reversible protein phosphoryla- tion is one of the major mechanisms used to control the function, localization and stability of proteins inside cells [1]. Therefore, the analysis of protein kinase activity and the phosphorylation level of their sub- strates are important for understanding signal trans- duction pathways at a molecular level. Many methods have been described for determining protein kinase activity and protein phosphorylation levels. In vitro, phosphorylation reactions can be monitored by incu- bating a protein kinase and a substrate in the presence of radioactive ATP ([ 32 P]ATP[cP]), followed by SDS ⁄ PAGE and autoradiography to quantify radio- activity in the substrate. When a peptide substrate is used, the amount of radioactivity incorporated into Keywords Cdc37; CK2; gel electrophoresis; Hsp90; protein kinase Correspondence Y. Miyata, Department of Cell & Developmental Biology, Graduate School of Biostudies, Kyoto University, Kitashirakawa Oiwake-cho, Sakyo-ku, Kyoto 606-8502, Japan Fax: +81 75 753 4235 Tel: +81 75 753 4231 E-mail: ymiyata@lif.kyoto-u.ac.jp (Received 22 March 2007, revised 9 August 2007, accepted 4 September 2007) doi:10.1111/j.1742-4658.2007.06090.x The CK2-dependent phosphorylation of Ser13 in cell division cycle pro- tein 37 (Cdc37), a kinase-specific heat shock protein 90 (Hsp90) cochaper- one, has previously been reported to be essential for the association of Cdc37 with signaling protein kinases [Bandhakavi S, McCann RO, Hanna DE & Glover CVC (2003) J Biol Chem 278, 2829–2836; Shao J, Prince T, Hartson SD & Matts RL (2003) J Biol Chem 278, 38117–38220; Miyata Y & Nishida E (2004) Mol Cell Biol 24, 4065–4074]. Here we describe a new phospho-specific antibody against Cdc37 that recognizes recombinant puri- fied Cdc37 only when incubated with CK2 in the presence of Mg 2+ and ATP. The replacement of Ser13 in Cdc37 by nonphosphorylatable amino acids abolished binding to this antibody. The antibody was specific for phosphorylated Cdc37 and did not crossreact with other CK2 substrates such as Hsp90 and FK506-binding protein 52. Using this antibody, we showed that complexes of Hsp90 with its client signaling kinases, Cdk4, MOK, v-Src, and Raf1, contained the CK2-phosphorylated form of Cdc37 in vivo. Immunofluorescent staining showed that Hsp90 and the phosphory- lated form of Cdc37 accumulated in epidermal growth factor-induced mem- brane ruffles. We further characterized the phosphorylation of Cdc37 using phospho-affinity gel electrophoresis. Our analyses demonstrated that the CK2-dependent phosphorylation of Cdc37 on Ser13 caused a specific gel mobility shift, and that Cdc37 in the complexes between Hsp90 and its cli- ent signaling protein kinases was in the phosphorylated form. Our results show the physiological importance of CK2-dependent Cdc37 phosphoryla- tion and the usefulness of phospho-affinity gel electrophoresis The light-harvesting antenna of the diatom Phaeodactylum tricornutum Evidence for a diadinoxanthin-binding subcomplex Ge ´ rard Guglielmi, Johann Lavaud*, Bernard Rousseau, Anne-Lise Etienne, Jean Houmard and Alexander V. Ruban† Organismes Photosynthe ´ tiques et Environnement, CNRS, De ´ partement de Biologie, Ecole Normale Supe ´ rieure, Paris, France Diatoms constitute a dominant group of phytoplank- tonic algae, which play an important role in the car- bon, silica and nitrogen biogeochemical cycles [1–3]. Their photosynthetic efficiency and subsequent produc- tivity depend upon the light environment, which can vary greatly as a result of water motion [4,5]. Fluctu- ating irradiances and, especially, excess light exposure can be harmful for photosynthesis, in particular photo- system II (PSII), causing a decrease in productivity and fitness [6,7]. One of the photoprotective mecha- nisms used by diatoms is the dissipation of excess energy in the light-harvesting complex (LHC) of PSII to prevent overexcitation of the photosystems [the so-called nonphotochemical chlorophyll fluorescence Keywords diatom; fucoxanthin; light-harvesting complex; photoprotection; xanthophyll cycle Correspondence G. Guglielmi, Organismes Photo- synthe ´ tiques et Environnement, UMR 8541 CNRS, De ´ partement de Biologie, Ecole Normale Supe ´ rieure, 46 rue d’Ulm, 75230 Paris cedex 05, France Fax: +33 1 44 32 39 41 Tel: +33 1 44 32 35 30 E-mail: ggugliel@biologie.ens.fr Present address *Pflanzliche O ¨ kophysiologie, Fachbereich Biologie, Universita ¨ t Konstanz, Germany †The Robert Hill Institute, Department of Molecular Biology and Biotechnology, University of Sheffield, UK Note A website is available: http://www.biologie. ens.fr/opeaec/ (Received 31 May 2005, revised 1 July 2005, accepted 5 July 2005) doi:10.1111/j.1742-4658.2005.04846.x Diatoms differ from higher plants by their antenna system, in terms of both polypeptide and pigment contents. A rapid isolation procedure was designed for the membrane-intrinsic light harvesting complexes (LHC) of the diatom Phaeodactylum tricornutum to establish whether different LHC subcomplexes exist, as well to determine an uneven distribution between them of pigments and polypeptides. Two distinct fractions were separated that contain functional oligomeric complexes. The major and more stable complex ( 75% of total polypeptides) carries most of the chlorophyll a, and almost only one type of carotenoid, fucoxanthin. The minor complex, carrying  10–15% of the total antenna chlorophyll and only a little chlorophyll c, is highly enriched in diadinoxanthin, the main xanthophyll cycle carotenoid. The two complexes also differ in their polypeptide com- position, suggesting specialized functions within the antenna. The diadinoxanthin-enriched complex could be where the de-epoxidation of diadinoxanthin into diatoxanthin mostly occurs. Abbreviations a-DM, n-dodecyl-a, D-maltoside; b-DM, n-dodecyl-b,D-maltoside; CAB protein, chlorophyll a-binding protein; Chl, chlorophyll; DD, diadinoxanthin; DT, diatoxanthin; FCP, fucoxanthin chlorophyll proteins; LHC, light harvesting complex; NPQ, nonphotochemical chlorophyll fluorescence quenching; PSI, photosystem I; PSII, photosystem II; XC, xanthophyll cycle. FEBS Journal 272 (2005) 4339–4348 ª 2005 FEBS 4339 quenching (NPQ)]. NPQ is triggered by a trans- thylakoidal proton gradient (DpH) and results from a modulation in the xanthophyll content [8–12]. In dia- toms, the xanthophyll cycle (XC) is made up of diadino- xanthin (DD), which is converted under an excess of light into its de-epoxidized form, diatoxanthin (DT) [13]. ... ([link]) 3/12 The Light- Dependent Reactions of Photosynthesis The colors of visible light not carry the same amount of energy Violet has the shortest wavelength and therefore carries the most energy,... frequencies of radiation ([link]) The difference between wavelengths relates to the amount of energy carried by them 2/12 The Light- Dependent Reactions of Photosynthesis The sun emits energy in the form... photosynthesis within a leaf Section Summary The pigments of the first part of photosynthesis, the light- dependent reactions, absorb energy from sunlight A photon strikes the antenna pigments of photosystem