Conformations of QB-. trapped by B side electron transfer in reaction centers from Rhodobacter sphaeroides (vol 86, pg 11a, 2004)

The electron transfer to Q(B) in the native RC has been proposed to be limited by a conformational gating step (1). We have constructed a quintuple mutant RC that allows direct reduction of Q(B) by bactcriopheohytin via the normally inactive B-branch utilizing a greater driving force for electron transfer in samples lacking active Q(A). This mutant consists of 5 amino acid replacements - one to displace Q(A) (Ala-M260->Trp) (2) three to inhibit A-side transfer to BChlA (Gly-M203->Asp, Tyr-M210-> Phe, Leu-M214->His) and one to promote B-side transfer (Phe-L181->Tyr). RCs frozen in the dark and illuminated at 80 K showed a light induced EPR signal of D+.Q(B)(-). in -10% of the RC sample. The lack of a narrow Q(B)(-). signal indicates a magnetic interaction between QB- and the Fe2+. These results suggest that similar to10% of the RCs have the quinone in the more thermodynamically stable proximal site (3). The quantum yield for formation was similar to7%, similar to those previously reported for other B-side mutants (4,5). The lifetime of the dark generated D+.Q(B)(-). is tau similar to 3s at 80 K. This result is in contrast to PCs frozen in the light, in which similar to100% D+.Q(B)(-), was formed with a lifetime > 10(5)s (6). The large difference in stability is attributed to an undetermined conformational change that occurs only at higher temperatures and stabilizes the reduced form of Q(B). This change may be responsible for the conformational gate. *Supported by NIH.