Similarities and differences between the characteristics of gibberellin-binding protein and gibberellin 2-oxidases in adzuki bean (Vigna angularis) seedlings.
Gibberellin-binding proteins (GBPs) were purified ca. 230,000 fold. The characteristics of adzuki GBP were examined and compared with those of a recombinant gibberellin 2-oxidase (rVaGA2oxA1) that was fused with glutathione S-transferase (GST). VaGA2oxA1 was most abundantly expressed in etiolated adzuki bean seedlings, and VaGA2oxA1 and GBPs from adzuki bean seedlings showed gibberellin-binding activity when incubated with 2-oxoglutarate and Co2+. Both rVaGA2oxA1 and partially purified GBPs from adzuki bean seedlings showed very similar selectivity to gibberellins in binding assays, where biologically active gibberellins such as GA4, GA3, GA7, and GA1 showed higher binding affinity than biologically inactive gibberellins such as GA8, GA34, and 3-epi-GA4. The polyclonal antibody raised against rVaGA2oxA1 cross-reacted with all rVaGA2oxs (rVaGA2oxA1, rVaGA2oxA2, rVaGA2oxB1, rVaGA2oxB2, and rVaGA2oxB3) whose cDNAs were cloned from adzuki bean seedlings. Treated with the antibody, the recombinants that originally showed gibberellin-binding activity lost both binding activity and enzymatic activity. In contrast to the recombinants, the gibberellin-binding activity of GBPs from adzuki bean seedlings was hardly affected by the antibody treatment. The GBPs showed very weak gibberellin 2-oxidase-like activity, and it was not affected by the antibody treatment either. These observations suggest that a major component that showed GA-binding activity was apparently different from any gibberellin 2-oxidase cloned from the seedlings.
Park
Seung-Hyun
SH
Department of Applied Biological Chemistry, University of Tokyo, Japan.
Nakajima
Masatoshi
M
Hasegawa
Morifumi
M
Yamaguchi
Isomaro
I
D002352:Carrier Proteins / Q000302:isolation & purification / Q000378:metabolism*; D002853:Chromatography, Liquid; D007887:Fabaceae / Q000201:enzymology / Q000254:growth & development / Q000378:metabolism*; D006899:Mixed Function Oxygenases / Q000302:isolation & purification / Q000378:metabolism*; D010750:Phosphoproteins / Q000302:isolation & purification / Q000378:metabolism*; D010940:Plant Proteins / Q000302:isolation & purification / Q000378:metabolism*; D036226:Seedlings / Q000201:enzymology / Q000378:metabolism*
D002352:Carrier Proteins; D010750:Phosphoproteins; D010940:Plant Proteins; C099131:gibberellin-binding phosphoprotein, plant; D006899:Mixed Function Oxygenases; C121463:gibberellin 2-dioxygenase