Polarization-modulated infrared spectroscopy and X-ray reflectivity of photosystem II core complex at the gas-water interface

The state of photosystem II core complex (PS II CC) in monolayer at the gas -water interface was investigated using in situ polarization-nodulated infr ared reflection absorption spectroscopy and x-ray reflectivity techniques. Two approaches for preparing and manipulating the monolayers were examined and compared. In the first, PS II CC was compressed immediately after sprea ding at an initial surface pressure of 5.7 mN/m, whereas in the second, the monolayer was incubated for 30 min at an initial surface pressure of 0.6 m N/m before compression, in the first approach, the protein complex maintain ed its native alpha-helical conformation upon compression, and the secondar y structure of PS II CC was found to be stable for 2 h. The second approach resulted in films showing stable surface pressure below 30 mN/m and the pr esence of large amounts of beta-sheets, which indicated denaturation of PS II CC. Above 30 mN/m, those films suffered surface pressure instability, wh ich had to be compensated by continuous compression. This instability was c orrelated with the formation of new alpha-helices in the film. Measurements at 4 degrees C strongly reduced denaturation of PS II CC. The x-ray reflec tivity studies indicated that the spread film consists of a single protein layer at the gas-water interface. Altogether, this study provides direct st ructural and molecular information on membrane proteins when spread in mono layers at the gas-water interface.