Document Detail

NAD turnover and utilisation of metabolites for RNA synthesis in a reaction sensing the redox state of the cytochrome b6f complex in isolated chloroplasts.
MedLine Citation:
PMID:  7505745     Owner:  NLM     Status:  MEDLINE    
NAD is normally regarded as a redox molecule or as the substrate for ADP-ribosylation reactions. In this study, we describe the rapid metabolism of NAD by Percoll-gradient-purified lettuce chloroplasts and show that the adenine moiety can be incorporated into RNA in a dark-activated reaction that senses the redox state of the cytochrome b6f complex. Isolated chloroplasts rapidly metabolised radiolabelled NAD+ to 5'-AMP (within seconds) and adenosine during a 60-min incubation in vitro; the products were analysed by high-performance liquid chromatography. No radiolabelled ADP-ribose was detected. Radioactivity was incorporated into trichloroacetic-acid-insoluble material during this period, with approximately 2-4-fold more incorporation occurring in the dark. Most of this radiolabel was rendered acid-soluble by dilute alkaline digestion at 37 degrees C, yielding an approximately equal mixture of 2'-AMP and 3'-AMP, and by RNase digestion, identifying the acid-insoluble radioactive material as RNA. Protein-bound ADP-ribose would have yielded 5'-AMP and/or oligomeric/polymeric ADP-ribose after alkali digestion. The utilisation of NAD metabolites for RNA synthesis was restricted to the thylakoid compartment of the chloroplast. The use of a variety of electron-transport inhibitors such as 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone, bromanil (tetrabromo-1,4-benzoquinone), electron donors (dithiothreitol), electron acceptors (ferricyanide) and an uncoupler showed that the incorporation of radiolabel from NAD into acid-insoluble material was favoured when the cytochrome b6f complex was in the oxidised state (as pertaining to incubations in the dark).
C K Pearson; S B Wilson; R Schaffer; A W Ross
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  European journal of biochemistry / FEBS     Volume:  218     ISSN:  0014-2956     ISO Abbreviation:  Eur. J. Biochem.     Publication Date:  1993 Dec 
Date Detail:
Created Date:  1994-02-03     Completed Date:  1994-02-03     Revised Date:  2007-07-23    
Medline Journal Info:
Nlm Unique ID:  0107600     Medline TA:  Eur J Biochem     Country:  GERMANY    
Other Details:
Languages:  eng     Pagination:  397-404     Citation Subset:  IM    
Department of Molecular and Cell Biology, University of Aberdeen, Scotland.
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MeSH Terms
Chloroplasts / enzymology*
Chromatography, High Pressure Liquid
Cytochromes / chemistry,  metabolism*
Cytochromes f
Electron Transport / drug effects
Hydrogen-Ion Concentration
NAD / metabolism*
RNA / biosynthesis*
Reg. No./Substance:
0/Cytochromes; 0/Radioisotopes; 53-84-9/NAD; 63231-63-0/RNA; 9035-46-5/Cytochromes f

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