Cloning, sequence analysis and expression in Escherichia coli of the gene encoding a uricase from the yeast-like symbiont of the brown planthopper, Nilaparvata lugens.
A urate oxidase (uricase; EC 1.7.3.3) gene of the yeast-like fungal endosymbiont of the brown planthopper, Nilaparvata lugens, was cloned, and sequenced together with its flanking regions. The gene comprised a open reading frame of 987 bp, that was split into two parts by a single 96 bp intron. The encoded uricase was 296 amino acids with 62% sequence identity with that of Aspergillus flavus. The molecular weight deduced was 32,882, and the predicted isoelectric point was 6.06. The symbiont's uricase conserved all the known consensus motifs, except the C-terminal PTS-1, Ser-basic-Leu. The leucine at the third position of PTS-1 was replaced by serine in the C-terminus of the symbiont's uricase. The symbiont's uricase gene was successfully expressed in Escherichia coli, and the product, tagged with histidine residues, was purified. The symbiont's uricase, thus produced, was as active as those from plants and animals, but less active than those from other fungi.
Hongoh
Y
Y
Department of Biological Sciences, Graduate School of Science, University of Tokyo, Japan. ss67243@hongo.ecc.u-tokyo.ac.jp
Sasaki
T
T
Ishikawa
H
H
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D004271:DNA, Fungal / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics*; D005658:Fungi / Q000201:enzymology* / Q000235:genetics* / Q000302:isolation & purification; D015870:Gene Expression; D005800:Genes, Fungal; D006430:Hemiptera / Q000382:microbiology*; D007526:Isoelectric Point; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017386:Sequence Homology, Amino Acid; D013559:Symbiosis; D014503:Urate Oxidase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
D017931:DNA Primers; D018076:DNA, Complementary; D004271:DNA, Fungal; D014503:Urate Oxidase