AN ATTEMPT TO INCREASE THERMOSTABILITY OF THE MUTANT PHOTOSYNTHETIC REACTION CENTER OF Cereibacter sphaeroides USING DISULFIDE BONDS

Methods of site-directed mutagenesis are successfully used in structural and functional studies of photosynthetic reaction centers (RCs). It has been noted that many mutations near electron transfer cofactors reduce the temperature stability of Cereibacter sphaeroides RCs and affect the number of RCs in membranes. We previously reported [Selikhanov et al. (2023) Membranes, 25, 154] that the introduction of inter-subunit disulfide bridges on the periplasmic or cytoplasmic surface of the complex promotes an increase in the thermal stability of C. sphaeroides RCs. In this work, an attempt was made to increase the thermal stability of the mutant RC with the Ile M206 – Gln substitution by introducing inter-subunit disulfide bonds. This RC is of considerable interest for studying the mechanisms of early electron transfer processes in RCs. The effect of mutations on the amount of RCs in chromatophores was analyzed and it was found that the I(M206)Q mutation leads to a twofold decrease in the RC content in chromatophores, the introduction of disulfide bonds on the cytoplasmic or periplasmic sides of the complex reduces the amount of RCs in membranes by a third, the triple substitution I(M206)Q/G(M19)C/T(L214)C – almost 4 times, and substitutions I(M206)Q/V(M84)C/G(L278)C lead to disruption of RC assembly in the membrane. It was shown that the introduction of an inter-subunit S-S bond on the cytoplasmic surface of the complex did not have a significant effect on the thermal stability of the I(M206)Q RC. Our own and literature data on the factors influencing the assembly processes and ensuring the stability of the structure of integral membrane complexes are discussed.