Identification and characterization of a heat-induced isoform of aldolase in oat chloroplast.
An analysis of protein synthesis at elevated temperatures in oat (Avena sativa) leaves revealed a heat-induced 44 kDa polypeptide. A cDNA library of heat-treated leaves was constructed and screened with specific antibodies raised against this 44 kDa polypeptide. A clone encoding the 44 kDa protein was identified as a form of the chloroplast-localized fructose-bisphosphate aldolase (EC 4.1.2.13). Northern and western blot analyses indicated heat-induced accumulation of the chloroplast aldolase isoform at both the RNA and protein level. Heat inducibility was restricted to the chloroplastic form of the enzyme, and was not observed for the cytoplasmic aldolase. The heat-induced isoform co-purified with thykaloid fractions, as confirmed by immunoassay and activity analyses. However, when thylakoid membranes were treated with proteinase K, the aldolase isoform completely disappeared, suggesting that this enzyme is not embedded but rather tends to adhere to the chloroplast membranes. Immunoblot analysis of other plant species revealed similar heat induction of thykaloid-associated aldolase homologues, suggesting the possible existence of a universal control mechanism for this enzyme's heat tolerance
Michelis
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R
Faculty of Biology, Technion-Israel Institute of Technology, Haifa, Israel.
Gepstein
S
S
D000595:Amino Acid Sequence; D018554:Avena / Q000201:enzymology / Q000235:genetics*; D015152:Blotting, Northern; D002736:Chloroplasts / Q000201:enzymology*; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D015180:Electrophoresis, Gel, Two-Dimensional; D005634:Fructose-Bisphosphate Aldolase / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D018506:Gene Expression Regulation, Plant; D006358:Hot Temperature; D007527:Isoenzymes / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D008969:Molecular Sequence Data; D010944:Plants / Q000201:enzymology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D014018:Tissue Distribution
D018076:DNA, Complementary; D007527:Isoenzymes; D012333:RNA, Messenger; D005634:Fructose-Bisphosphate Aldolase