Light-induced changes in the distribution of the 36000-Mr polypeptide of NADPH-protochlorophyllide oxidoreductase within different cellular compartments of barley (Hordeum vulgare L.) : I. Localization by immunoblotting in isolated plastids and total leaf extracts.

Changes in the relative content of NADPH-protochlorophyllide oxidoreductase during the light-induced greening of barley plants were measured both in the total leaf extract as well as in intact and broken plastids. The enzyme protein was identified by its apparent molecular weight and its immunological crossreactivity with an antiserum directed against the NADPH-protochlorophyllide oxidoreductase. The monospecificity of the antiserum was tested by two different criteria: i. The antiserum was purified by affinity chromatography. ii. It was demonstrated that the antiserum crossreacts with only those polypeptides which appear to be enzymatically active. In the fraction of broken plastids isolated from leaves of briefly illuminated barley plants the concentration of the enzyme protein was reduced drastically. Our results indicate that this decrease in enzyme protein content is the consequence of an artificial proteolytic breakdown of the membrane-bound enzyme protein. In intact plastids and in the total leaf extract the concentration of the enzyme protein did not change dramatically during the first 4 to 6 h of illumination. However, when the exposure to continuous white light was extended further the concentration of the enzyme protein in intact plastids began to decline rapidly while in total leaf extracts the concentration remained almost constant for the next 10 h of light. These results indicate that part of the enzyme protein may be localized outside of the plastid compartment.