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Xanthophyll Cycle in the Light of Thylakoid Membrane Lipids - Membrane Packing, Curvature Elastic Stress and Enzyme Binding

Szilagyi, Anna LU (2007)
Abstract (Swedish)
Popular Abstract in Swedish

Fotosyntetiska membraner utsätts för stress vid höga ljusintensiteter och skador kan lätt uppstå. En av de viktigaste skyddande substanserna i växter är karotenoiden zeaxantin. Zeaxantin bildas från violaxantin i xantofyllcykeln och bildningen katalyseras av enzymet violaxantin de-epoxidas (VDE). I aktiv form är enzymet bundet till det fotosyntetiska membranet (tylakoidmembranet). Enzymet kräver lågt pH, askorbinsyra (vitamin C) och en speciell lipid, monogalactosyldiacylglycerol (MGDG). Produkten zeaxantin har olika funktioner. Den viktigaste är att delta i skyddet av fotosyntesmaskineriet mot överskott av ljus. Bildningen av zeaxantin har också föreslagits kunna påverka rörligheten hos... (More)
Popular Abstract in Swedish

Fotosyntetiska membraner utsätts för stress vid höga ljusintensiteter och skador kan lätt uppstå. En av de viktigaste skyddande substanserna i växter är karotenoiden zeaxantin. Zeaxantin bildas från violaxantin i xantofyllcykeln och bildningen katalyseras av enzymet violaxantin de-epoxidas (VDE). I aktiv form är enzymet bundet till det fotosyntetiska membranet (tylakoidmembranet). Enzymet kräver lågt pH, askorbinsyra (vitamin C) och en speciell lipid, monogalactosyldiacylglycerol (MGDG). Produkten zeaxantin har olika funktioner. Den viktigaste är att delta i skyddet av fotosyntesmaskineriet mot överskott av ljus. Bildningen av zeaxantin har också föreslagits kunna påverka rörligheten hos komponenter i tylakoidmembranet och är en av de centrala frågorna som jag behandlar i denna avhandling.



VDE har molekylvikten 43 kDa och är löslig i tylakoidernas inre (lumen) vid högt pH. Vid höga ljusintensiteter och därigenom mycket aktiv fotosyntes sjunker pH i lumen, VDE ändrar sin form och binder till membranet. Bindningen till membranet är avgörande för aktiviteten eftersom violaxantin befinner sig i membranet. I arbete I visar vi, med hjälp av riktad mutagenes, att fyra konserverade hisitidiner i VDE är väsentliga för det starka pH beroende enzymet har för sin inbindning till membranet.



I arbete II har vi studerat hur aktiviteten hos VDE påverkas av lipidernas packning i tylakoidmembranet. MGDG är en lipid som krävs för enzymets aktivitet men dess närvaro i membranet leder till inre spänningar i membranet. Genom att tillsätta substanser som ökar eller minskar spänningarna i membranet har vi kraftigt påverkat graden av omvandling av violaxantin. Ju mer spänningar desto mer violaxantin gick att omvandla. Vår slutsats är att VDE kräver spänningar i membranet för att fungera. Zeaxantin med sin utökade hydrofoba längd föreslogs göra membranet tätare och hjälpa till att minska spänningarna.



I arbete III visar vi för första gången att packning och rörlighet i tylakoidmembran kan studeras med hjälp av en fluorescerande substans, laurdan. Laurdan ändrar sina fluorescensegenskaper när dess egen och närmaste vattenmolekylers rörelse minskar. I detta arbete har vi kunnat visa att violaxantin, zeaxantin och ?-tocopherol alla gör membranet mindre rörligt men att zeaxantin är effektivast. Omvandlingen av violaxantin kan nu ses i ett vidare perspektiv. Eftersom växter ofta utsätts för värme samtidigt som de utsätts för hög ljusintesitet ger bildningen av zeaxantin inte bara skydd av fotosyntesmaskineriet utan kan också hjälpa till att stabilisera membranstrukturen. (Less)
Abstract
The xanthophyll cycle involves the light-dependent and reversible conversion of violaxanthin to zeaxanthin. The enzyme, violaxanthin de-epoxidase (VDE) catalyses the conversion of violaxanthin to zeaxanthin, via the intermediate antheraxanthin. VDE is a membrane-hosted enzyme during its activity. Low lumenal pH, violaxanthin, ascorbate and monogalactosyldiacylglycerol are required for activity. The formed zeaxanthin is involved in the protection of the photosynthetic apparatus from overexcitation. Apart from the energy-dissipating role, the xanthophyll cycle has been associated with a modulation of the physical properties of the thylakoid membrane.



During photosynthetic activity, the lumenal pH drops and VDE undergoes a... (More)
The xanthophyll cycle involves the light-dependent and reversible conversion of violaxanthin to zeaxanthin. The enzyme, violaxanthin de-epoxidase (VDE) catalyses the conversion of violaxanthin to zeaxanthin, via the intermediate antheraxanthin. VDE is a membrane-hosted enzyme during its activity. Low lumenal pH, violaxanthin, ascorbate and monogalactosyldiacylglycerol are required for activity. The formed zeaxanthin is involved in the protection of the photosynthetic apparatus from overexcitation. Apart from the energy-dissipating role, the xanthophyll cycle has been associated with a modulation of the physical properties of the thylakoid membrane.



During photosynthetic activity, the lumenal pH drops and VDE undergoes a conformational change causing the active enzyme to bind to the lumenal side of the thylakoid membrane. Binding studies of VDE to the thylakoid membranes showed that the conserved histidine residues in the lipocalin region are involved in the conformational change of the enzyme.



By altering the ratio between the lamellar and non-lamellar structures in the thylakoid membrane, the activity of VDE was influenced. When the thylakoids were treated with linolenic acid which increases non-lamellar prone structures in the membrane, enhanced zeaxanthin formation was seen. A model is proposed involving membrane curvature stress, thylakoid membrane packing and the xanthophyll cycle. According to this model the different xanthophyll pigments have different preferences for different membrane structures and can themselves induce differentially curved membrane regions. Zeaxanthin with its increased molecular hydrophobic length affects lipid packing, and brings about a release of curvature stress leading to a less favoured lipid environment for VDE.



Xanthophylls, based on their rigid molecular structure, have been suggested to modify membrane structure, more specifically to decrease membrane fluidity. We have shown that accumulation of zeaxanthin in the isolated thylakoid membrane resulted in an increased rigidity of the membrane measured as a red-shifted fluorescence emission spectrum using laurdan, as a probe.



These results further support the hypothesis that xanthophyll cycle pigments are located in the lipid phase of the thylakoid membrane and affect membrane physical properties. (Less)
Please use this url to cite or link to this publication:
author
supervisor
opponent
  • Dr associate professor Funk, Christiane, Umeå universitet, Umeå
organization
publishing date
type
Thesis
publication status
published
subject
keywords
steroider, Lipids, steroids, membranes, Lipider, enzymologi, HPLC, laurdan, fluorescence spectroscopy, Proteins, enzymology, Proteiner, curvature elastic stress, enzyme binding, thylakoid membrane, membrane fluidity, lipid packing, histidine, VDE, zeaxanthin, xanthophyll cycle, mebran
pages
117 pages
publisher
Department of Biochemistry, Lund University
defense location
Kemicentrum, Lecture hall K:D, Getingevägen 60
defense date
2007-05-15 10:30
ISBN
978-91-7422-157-2
language
English
LU publication?
yes
id
dc1e6bc1-3bcf-4575-9160-7bcde2fab866 (old id 548507)
date added to LUP
2007-10-10 10:51:02
date last changed
2016-09-19 08:45:02
@phdthesis{dc1e6bc1-3bcf-4575-9160-7bcde2fab866,
  abstract     = {The xanthophyll cycle involves the light-dependent and reversible conversion of violaxanthin to zeaxanthin. The enzyme, violaxanthin de-epoxidase (VDE) catalyses the conversion of violaxanthin to zeaxanthin, via the intermediate antheraxanthin. VDE is a membrane-hosted enzyme during its activity. Low lumenal pH, violaxanthin, ascorbate and monogalactosyldiacylglycerol are required for activity. The formed zeaxanthin is involved in the protection of the photosynthetic apparatus from overexcitation. Apart from the energy-dissipating role, the xanthophyll cycle has been associated with a modulation of the physical properties of the thylakoid membrane.<br/><br>
<br/><br>
During photosynthetic activity, the lumenal pH drops and VDE undergoes a conformational change causing the active enzyme to bind to the lumenal side of the thylakoid membrane. Binding studies of VDE to the thylakoid membranes showed that the conserved histidine residues in the lipocalin region are involved in the conformational change of the enzyme.<br/><br>
<br/><br>
By altering the ratio between the lamellar and non-lamellar structures in the thylakoid membrane, the activity of VDE was influenced. When the thylakoids were treated with linolenic acid which increases non-lamellar prone structures in the membrane, enhanced zeaxanthin formation was seen. A model is proposed involving membrane curvature stress, thylakoid membrane packing and the xanthophyll cycle. According to this model the different xanthophyll pigments have different preferences for different membrane structures and can themselves induce differentially curved membrane regions. Zeaxanthin with its increased molecular hydrophobic length affects lipid packing, and brings about a release of curvature stress leading to a less favoured lipid environment for VDE.<br/><br>
<br/><br>
Xanthophylls, based on their rigid molecular structure, have been suggested to modify membrane structure, more specifically to decrease membrane fluidity. We have shown that accumulation of zeaxanthin in the isolated thylakoid membrane resulted in an increased rigidity of the membrane measured as a red-shifted fluorescence emission spectrum using laurdan, as a probe.<br/><br>
<br/><br>
These results further support the hypothesis that xanthophyll cycle pigments are located in the lipid phase of the thylakoid membrane and affect membrane physical properties.},
  author       = {Szilagyi, Anna},
  isbn         = {978-91-7422-157-2},
  keyword      = {steroider,Lipids,steroids,membranes,Lipider,enzymologi,HPLC,laurdan,fluorescence spectroscopy,Proteins,enzymology,Proteiner,curvature elastic stress,enzyme binding,thylakoid membrane,membrane fluidity,lipid packing,histidine,VDE,zeaxanthin,xanthophyll cycle,mebran},
  language     = {eng},
  pages        = {117},
  publisher    = {Department of Biochemistry, Lund University},
  school       = {Lund University},
  title        = {Xanthophyll Cycle in the Light of Thylakoid Membrane Lipids - Membrane Packing, Curvature Elastic Stress and Enzyme Binding},
  year         = {2007},
}