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Isolation, sequencing and expression of cDNA sequences encoding uroporphyrinogen decarboxylase from tobacco and barley

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Abstract

We have cloned and sequenced a full-length cDNA for uroporphyrinogen decarboxylase (UROD, EC 4.1.1.37) from tobacco (Nicotiana tabacum L.) and a partial cDNA clone from barley (Hordeum vulgare L.). The cDNA of tobacco encodes a protein of 43 kDa, which has 33% overall similarity to UROD sequences determined from other organisms. We propose that tobacco UROD has an N-terminal extension of 39 amino acid residues. This extension is most likely a chloroplast transit sequence. The in vitro translation product of UROD was imported into pea chloroplasts and processed to ca. 39 kDa. A truncated cDNA, from which the putative transit peptide had been deleted, was used to over-express the mature UROD in Escherichia coli. Purified protein showed UROD activity, thus providing an adequate source for subsequent enzymatic characterization and inhibition studies. Expression of UROD was investigated by northern and western blot analysis during greening of etiolated barley seedlings, and in segments of barley primary leaves grown under day/night cycles. The amount of RNA and protein increased during illumination Maximum UROD-RNA levels were detected in the basal segments relative to the top of the leaf.

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Abbreviations

ALA:

5-aminolevulinic acid

copro:

coproporphyrin

coprogen:

coproporphyrinogen

protogen IX:

protoporphyrinogen IX

UROD:

uroporphyrinogen decarboxylase

uro:

uroporphyrin

urogen:

uroporphyrinogen

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  1. Livio Trainotti

    Present address: Department of Biology, University of Padova, Via Trieste 75, 35121, Padova, Italy

Authors and Affiliations

  1. Institute of plant genetics and crop plant research, Corrensstraße 3, D-06466, Gatersleben, Germany

    Hans-Peter Mock, Livio Trainotti, Elisabeth Kruse & Bernhard Grimm

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  1. Hans-Peter Mock
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  4. Bernhard Grimm
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Mock, HP., Trainotti, L., Kruse, E. et al. Isolation, sequencing and expression of cDNA sequences encoding uroporphyrinogen decarboxylase from tobacco and barley. Plant Mol Biol 28, 245–256 (1995). https://doi.org/10.1007/BF00020244

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