Molecular cloning and characterization of a maize transglutaminase complementary DNA.
Two related complementary DNA clones, TGZ15 and TGZ21, encoding active maize transglutaminase (TGase) have been isolated for the first time in plants by molecular cloning (Patent Pending PCT/ES03/00247). Southern and northern blot analyses indicate that the two cDNAs probably corresponded to two different single-copy genes in the maize genome. Northern blot analyses revealed that the transcript is expressed preferentially in young leaves and differentiated embryogenic maize callus. This expression is dependent on light exposure time. TGase activity of the proteins encoded by clones TGZ15 and TGZ21 was detected in bacterial extracts overexpressing them, using two enzymatic assays. TGase activity was significantly higher than that of the empty-phagemid bacterial extracts. As in other TGases, this activity was inhibited by monodansyl cadaverine (MDC), GTP and the absence of exogenous Ca(2+). Likewise, light-stimulated Ca(2+)-dependent TGase activity was detected in thylakoids and grana of maize chloroplast, which was inhibited by MDC, GTP, DIECA and Diuron.
Villalobos
E
E
Departament de Genètica Molecular, Institut de Biología Molecular de Barcelona, Cordi Girona 18-26, 08034 Barcelona, Spain.
Santos
M
M
Talavera
D
D
Rodríguez-Falcón
M
M
Torné
J M
JM
D000595:Amino Acid Sequence; D001483:Base Sequence; D001710:Biotin / Q000378:metabolism; D015152:Blotting, Northern; D015153:Blotting, Western; D002103:Cadaverine / Q000378:metabolism; D020134:Catalytic Domain / Q000235:genetics; D002736:Chloroplasts / Q000201:enzymology / Q000378:metabolism; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics*; D015971:Gene Expression Regulation, Enzymologic; D018506:Gene Expression Regulation, Plant; D018745:Genome, Plant; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D010802:Phylogeny; D010936:Plant Extracts / Q000378:metabolism; D021381:Protein Transport; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid; D020524:Thylakoids / Q000201:enzymology; D011503:Transglutaminases / Q000235:genetics* / Q000378:metabolism; D003313:Zea mays / Q000201:enzymology / Q000235:genetics*
D018076:DNA, Complementary; D007527:Isoenzymes; D010936:Plant Extracts; D011994:Recombinant Proteins; D001710:Biotin; D011503:Transglutaminases; D002103:Cadaverine