Figure 8. Tryptic digestion of phosphorylated NDH-F polypeptide in the immunopurified Ndh complex.

Ndh complex (0.5 μg) was immunopurified from freshly prepared thylakoids and incubated in 100 mM Tris/HCl (pH 7.5) and 10 units of trypsin at 20 °C. At the indicated times, reactions were stopped by the addition of soya-bean trypsin inhibitor [75 ng·(μg of trypsin)⁻¹] in SDS sample buffer and then boiling for 10 min. Samples were subjected to Tricine-SDS/PAGE (10% acrylamide) and Western-blot analysis using a monospecific NDH-F antiserum (A) or a monoclonal phosphothreonine antibody (B).