The molybdenum cofactor biosynthetic protein Cnx1 complements molybdate-repairable mutants, transfers molybdenum to the metal binding pterin, and is associated with the cytoskeleton.
Molybdenum (Mo) plays an essential role in the active site of all eukaryotic Mo-containing enzymes. In plants, Mo enzymes are important for nitrate assimilation, phytohormone synthesis, and purine catabolism. Mo is bound to a unique metal binding pterin (molybdopterin [MPT]), thereby forming the active Mo cofactor (Moco), which is highly conserved in eukaryotes, eubacteria, and archaebacteria. Here, we describe the function of the two-domain protein Cnx1 from Arabidopsis in the final step of Moco biosynthesis. Cnx1 is constitutively expressed in all organs and in plants grown on different nitrogen sources. Mo-repairable cnxA mutants from Nicotiana plumbaginifolia accumulate MPT and show altered Cnx1 expression. Transformation of cnxA mutants and the corresponding Arabidopsis chl-6 mutant with cnx1 cDNA resulted in functional reconstitution of their Moco deficiency. We also identified a point mutation in the Cnx1 E domain of Arabidopsis chl-6 that causes the molybdate-repairable phenotype. Recombinant Cnx1 protein is capable of synthesizing Moco. The G domain binds and activates MPT, whereas the E domain is essential for activating Mo. In addition, Cnx1 binds to the cytoskeleton in the same way that its mammalian homolog gephyrin does in neuronal cells, which suggests a hypothetical model for anchoring the Moco-synthetic machinery by Cnx1 in plant cells.
Schwarz
G
G
Botanical Institute, Technical University of Braunschweig, 38023 Braunschweig, Germany.
Schulze
J
J
Bittner
F
F
Eilers
T
T
Kuper
J
J
Bollmann
G
G
Nerlich
A
A
Brinkmann
H
H
Mendel
R R
RR
D000595:Amino Acid Sequence; D000818:Animals; D017360:Arabidopsis / Q000235:genetics; D029681:Arabidopsis Proteins; D001692:Biological Transport; D037281:Calnexin; D003599:Cytoskeleton / Q000378:metabolism*; D005816:Genetic Complementation Test; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D008982:Molybdenum / Q000378:metabolism*; D009154:Mutation; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010947:Plants, Toxic; D011622:Pterins / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D014026:Nicotiana / Q000235:genetics
D029681:Arabidopsis Proteins; C080351:CNX1 protein, Arabidopsis; D008565:Membrane Proteins; D010940:Plant Proteins; D011622:Pterins; D037281:Calnexin; D008982:Molybdenum