In vitro-mutagenesis of NADPH:protochlorophyllide oxidoreductase B: two distinctive protochlorophyllide binding sites participate in enzyme catalysis and assembly.
NADPH:protochlorophyllide oxidoreductase (POR) B is a key enzyme for the light-induced greening of etiolated angiosperm plants. It is nucleus-encoded, imported into the plastids posttranslationally, and assembled into larger light-harvesting POR:protochlorophyllide complexes termed LHPP (Reinbothe et al., Nature 397:80-84, 1999). An in vitro-mutagenesis approach was taken to study the role of the evolutionarily conserved Cys residues in pigment binding. Four Cys residues are present in the PORB of which two, Cys276 and Cys303, established distinct pigment binding sites, as shown by biochemical tests, protein import studies, and in vitro-reconstitution experiments. While Cys276 constituted the Pchlide binding site in the active site of the enzyme, Cys303 established a second, low affinity pigment binding site that was involved in the assembly and stabilization of imported PORB enzyme inside etioplasts.
Reinbothe
Christiane
C
Lehrstuhl für Pflanzenphysiologie, Universität Bayreuth, Universitätsstrasse 30, 95447, Bayreuth, Germany. christiane.reinbothe@uni-bayreuth.de
Buhr
Frank
F
Bartsch
Sandra
S
Desvignes
Claire
C
Quigley
Françoise
F
Pesey
Hélène
H
Reinbothe
Steffen
S
D000595:Amino Acid Sequence; D001483:Base Sequence; D002384:Catalysis; D017931:DNA Primers; D008969:Molecular Sequence Data; D016296:Mutagenesis; D044925:Oxidoreductases Acting on CH-CH Group Donors / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid
D017931:DNA Primers; D044925:Oxidoreductases Acting on CH-CH Group Donors; C023886:protochlorophyllide reductase