Molecular cloning and characterization of an amidase from Arabidopsis thaliana capable of converting indole-3-acetamide into the plant growth hormone, indole-3-acetic acid.
Acylamidohydrolases from higher plants have not been characterized or cloned so far. AtAMI1 is the first member of this enzyme family from a higher plant and was identified in the genome of Arabidopsis thaliana based on sequence homology with the catalytic-domain sequence of bacterial acylamidohydrolases, particularly those that exhibit indole-3-acetamide amidohydrolase activity. AtAMI1 polypeptide and mRNA are present in leaf tissues, as shown by immunoblotting and RT-PCR, respectively. AtAMI1 was expressed from its cDNA in enzymatically active form and exhibits substrate specificity for indole-3-acetamide, but also some activity against L-asparagine. The recombinant enzyme was characterized further. The results show that higher plants have acylamidohydrolases with properties similar to the enzymes of certain plant-associated bacteria such as Agrobacterium-, Pseudomonas- and Rhodococcus-species, in which these enzymes serve to synthesize the plant growth hormone, indole-3-acetic acid, utilized by the bacteria to colonize their host plants. As indole-3-acetamide is a native metabolite in Arabidopsis thaliana, it can no longer be ruled out that one pathway for the biosynthesis of indole-3-acetic acid involves indole-3-acetamide-hydrolysis by AtAMI1.
Pollmann
Stephan
S
Lehrstuhl für Pflanzenphysiologie, Ruhr-Universität Bochum, D-44780 Bochum, Germany.
Neu
Daniel
D
Weiler
Elmar W
EW
D000581:Amidohydrolases / Q000235:genetics* / Q000378:metabolism*; D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000201:enzymology* / Q000235:genetics*; D003001:Cloning, Molecular; D015151:Immunoblotting; D007210:Indoleacetic Acids / Q000378:metabolism*; D008969:Molecular Sequence Data; D010937:Plant Growth Regulators / Q000096:biosynthesis*; D018515:Plant Leaves / Q000378:metabolism; D011549:Pseudomonas / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012231:Rhizobium / Q000235:genetics; D012240:Rhodococcus / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D007210:Indoleacetic Acids; D010937:Plant Growth Regulators; D012333:RNA, Messenger; D011994:Recombinant Proteins; C030737:indoleacetic acid; C015950:indoleacetamide; D000581:Amidohydrolases