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Open AccessArticle

Aerobic Barley Mg-protoporphyrin IX Monomethyl Ester Cyclase is Powered by Electrons from Ferredoxin

1
Department of Biology, Lund University, Sölvegatan 35B, 22362 Lund, Sweden
2
Faculty of Agriculture, Cairo University, Giza 12613, Egypt
3
Department of Food Science, University of Copenhagen, Rolighedsvej 26, DK-1958 Frederiksberg, Denmark
4
Department of Biology, Illinois Wesleyan University, Bloomington, IL P.O. Box 2900, USA
*
Author to whom correspondence should be addressed.
Plants 2020, 9(9), 1157; https://doi.org/10.3390/plants9091157
Received: 11 August 2020 / Revised: 3 September 2020 / Accepted: 4 September 2020 / Published: 8 September 2020
(This article belongs to the Collection Feature Papers in Plant Physiology and Metabolism)
Chlorophyll is the light-harvesting molecule central to the process of photosynthesis. Chlorophyll is synthesized through 15 enzymatic steps. Most of the reactions have been characterized using recombinant proteins. One exception is the formation of the isocyclic E-ring characteristic of chlorophylls. This reaction is catalyzed by the Mg-protoporphyrin IX monomethyl ester cyclase encoded by Xantha-l in barley (Hordeum vulgare L.). The Xantha-l gene product (XanL) is a membrane-bound diiron monooxygenase, which requires additional soluble and membrane-bound components for its activity. XanL has so far been impossible to produce as an active recombinant protein for in vitro assays, which is required for deeper biochemical and structural analyses. In the present work, we performed cyclase assays with soluble and membrane-bound fractions of barley etioplasts. Addition of antibodies raised against ferredoxin or ferredoxin-NADPH oxidoreductase (FNR) inhibited assays, strongly suggesting that reducing electrons for the cyclase reaction involves ferredoxin and FNR. We further developed a completely recombinant cyclase assay. Expression of active XanL required co-expression with an additional protein, Ycf54. In vitro cyclase activity was obtained with recombinant XanL in combination with ferredoxin and FNR. Our experiment demonstrates that the cyclase is a ferredoxin-dependent enzyme. Ferredoxin is part of the photosynthetic electron-transport chain, which suggests that the cyclase reaction might be connected to photosynthesis under light conditions. View Full-Text
Keywords: acsF; bchE; CHL27; chlorophyll biosynthesis; CRD1; FNR; Hordeum vulgare; XanL; Xantha-l acsF; bchE; CHL27; chlorophyll biosynthesis; CRD1; FNR; Hordeum vulgare; XanL; Xantha-l
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Stuart, D.; Sandström, M.; Youssef, H.M.; Zakhrabekova, S.; Jensen, P.E.; Bollivar, D.W.; Hansson, M. Aerobic Barley Mg-protoporphyrin IX Monomethyl Ester Cyclase is Powered by Electrons from Ferredoxin. Plants 2020, 9, 1157.

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