Regular paper Structure and activity of the Photosystem II manganese-stabilizing protein: role of the conserved disulfide bond

The 33-kDa manganese-stabilizing protein (MSP) of Photosystem II (PS II) maintains the functional stability of the Mn cluster in the enzyme’s active site. This protein has been shown to possess characteristics similar to those of the intrinsically disordered, or natively unfolded proteins [Lydakis-Simantiris et al. (1999b) Biochemistry 38: 404–414]. Alternately it was proposed that MSP should be classified as a molten globule, based in part on the hypothesis that its lone disulfide bridge is necessary for structural stability and function in solution [Shutova et al. (2000) FEBS Lett. 467: 137–140]. A site-directed mutant MSP (C28A,C51A) that eliminates the disulfide bond reconstitutes O2 evolution activity and binds to MSP-free PS II preparations at wild-type levels [Betts et al. (1996) Biochim. Biophys. Acta 1274: 135–142]. This mutant was further characterized by incubation at 90 C to determine the effect of loss of the disulfide bridge on MSP thermostability and solution structure. After heating at 90 C for 20 min, C28A,C51A MSP was still able to bind to PS II preparations at molar stoichiometries similar to those of WT MSP and reconstitute O2 evolution activity. A fraction of the protein aggregates upon heating, but after resolubili-zation, it regains the ability to bind to PS II and reconstitute O2 evolution activity. Characterization of the solution structure of C28A,C51A MSP, using CD spectroscopy, UV absorption spectroscopy, and ge