S. Engdahl et al., Association of the NADPH : protochlorophyllide oxidoreductase (POR) with isolated etioplast inner membranes from wheat, PLANT J, 27(4), 2001, pp. 297-304
Membrane association of NADPH:protochlorophyllide oxidoreductase (POR, EC:
1.6.99.1) with isolated prolamellar bodies (PLBs) and prothylakoids (PTs) f
rom wheat etioplasts was investigated. in vitro-expressed radiolabelled POR
, with or without transit peptide, was used to characterize membrane associ
ation conditions. Proper association of POR with PLBs and PTs did not requi
re the presequence, whereas NADPH and hydrolysable ATP were vital for the p
rocess. After treating the membranes with thermolysin, sodium hydroxide or
carbonate, a firm attachment of the POR protein to the membrane was found.
Although the PLBs and PTs differ significantly in their relative amount of
POR in vivo, no major differences in POR association capacity could be obse
rved between the two membrane systems when exogenous NADPH was added, Exper
iments run with only an endogenous NADPH source almost abolished associatio
n of POR with both PLBs and PTs. In addition, POR protein carrying a mutati
on in the putative nucleotide-binding site (ALA06) was unable to bind to th
e inner membranes in the presence of NADPH, which further demonstrates that
the co-factor is essential for proper membrane association. POR protein ca
rrying a mutation in the substrate-binding site (ALA24) showed less binding
to the membranes as compared to the wild type. The results presented here
introduce studies of a novel area of protein-membrane interaction, namely t
he association of proteins with a paracrystalline membrane structure, the P
LB.