Isolation of a gene encoding a glycosylated cytokinin oxidase from maize.
The major cytokinin oxidase in immature maize kernels was purified to homogeneity. Selected tryptic peptides were used to design degenerate oligonucleotide primers for PCR isolation of a fragment of the oxidase gene. Hybridization of the PCR fragment to a maize genomic library allowed isolation of a full-length cytokinin oxidase gene (ckx1). The gene encodes a protein of approximately 57 kDa that possesses a signal peptide, eight consensus N-glycosylation sequences and a consensus FAD binding sequence. Expression of ckx1 in Pichia caused secretion of active glycosylated cytokinin oxidase that contains a substrate-reducible FAD. The gene displays sequence homology with a putative oxidoreductase from Arabidopsis thaliana and with the fas5 gene from Rhodococcus fascians.
Biochemistry Department, University of Missouri, Columbia 65211, USA. morris@missouri.edu
R O
RO
Morris
K D
KD
Bilyeu
J G
JG
Laskey
N N
NN
Cheikh
D000595:Amino Acid Sequence; D003583:Cytokinins / Q000737:chemistry / Q000235:genetics; D017343:Genes, Plant; D006031:Glycosylation; D008969:Molecular Sequence Data; D010088:Oxidoreductases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid; D013379:Substrate Specificity; D003313:Zea mays / Q000201:enzymology* / Q000235:genetics*
D003583:Cytokinins; D010088:Oxidoreductases; C019984:cytokinin oxidase