The monofunctional cobalamin biosynthesis enzyme precorrin-3B synthase (CobZRR) is essential for anaerobic photosynthesis in Rhodospirillum rubrum but not for aerobic dark metabolism.

The in vivo physiological role of the gene cobZ, which encodes precorrin-3B synthase, which catalyzes the initial porphyrin ring contraction step of cobalamin biosynthesis via the cob pathway, has been demonstrated here for the first time. Cobalamin is known to be essential for an early step of bacteriochlorophyll biosynthesis in anoxygenic purple bacteria. The cobZ (cobZRR) gene of the purple bacterium Rhodospirillum rubrum was localized to a 23.5 kb insert of chromosomal DNA contained on the cosmid pSC4. pSC4 complemented several mutants of bacteriochlorophyll and carotenoid biosynthesis, due to the presence of the bchCX and crtCDEF genes at one end of the cosmid insert, flanking cobZRR. A second gene, citB/tcuB, immediately downstream of cobZRR, shows homologies to both a tricarballylate oxidoreductase (tcuB) and a gene (citB) involved in signal transduction during citrate uptake. CobZRR shows extensive homology to the N-terminal domain of the bifunctional CobZ from Rhodobacter capsulatus, and the R. rubrum citB/tcuB gene is homologous to the CobZ C-terminal domain. A mutant, SERGK25, containing a terminatorless kanamycin interposon inserted into cobZRR, could not grow by anaerobic photosynthesis, but grew normally under dark, aerobic and microaerophilic conditions with succinate and fructose as carbon sources. The anaerobic in vivo activity of CobZ indicates that it does not require oxygen as a substrate. The mutant excreted large amounts of protoporphyrin IX-monomethylester, a brown precursor of bacteriochlorophyll biosynthesis. The mutant was complemented either by the cobZRR gene in trans, or when exogenous cobalamin was added to the medium. A deletion mutant of tcuB/citB did not exhibit the cob phenotype. Thus, a role for tcuB/citB in cobalamin biosynthesis could not be confirmed.

Excitation energy pathways in the photosynthetic units of reaction center LM- and H-subunit deletion mutants of Rhodospirillum rubrum.

Light-induced reaction dynamics of isolated photosynthetic membranes obtained from wild-type (WT) and reaction center (RC)-subunit deletion strains SPUHK1 (an H-subunit deletion mutant) and SK Delta LM (an (L+M) deletion mutant) of the purple non-sulphur bacterium Rhodospirillum rubrum have been investigated by femtosecond transient absorption spectroscopy. Upon excitation of the spirilloxanthin (Spx) S(2) state at 546 nm, of the bacteriochlorophyll Soret band at 388 nm and probing spectral regions, which are characteristic for carotenoids, similar dynamics in the SPUHK1, SK Delta LM and WT strains could be observed. The excitation of Spx S(2) is followed by the simultaneous population of the lower singlet excited states S(1) and S* which decay with lifetimes of 1.4 and 5 ps, respectively for the mutants, and 1.4 and 4 ps, respectively, for the wild-type. The excitation of the BChl Soret band is followed by relaxation into BChl lower excited states which compete with excitation energy transfer BChl-to-Spx. The deexcitation pathway BChl(Soret) --> Spx(S(2)) --> Spx(S(1)) occurs with the same transition rate for all investigated samples (WT, SPUHK1 and SK Delta LM). The kinetic traces measured for the Spx S(1) --> S(N) transition display similar behaviour for all samples showing a positive signal which increases within the first 400 fs (i.e. the time needed for the excitation energy to reach the Spx S(1) excited state) and decays with a lifetime of about 1.5 ps. This suggests that the Spx excited state dynamics in the investigated complexes do not differ significantly. Moreover, a longer excited state lifetime of BChl for SPUHK1 in comparison to WT was observed, consistent with a photochemical quenching channel present in the presence of RC. For long delay times, photobleaching of the RC special pair and an electrochromic blue shift of the monomeric BChl a can be observed only for the WT but not for the mutants. The close similarity of the excited state decay processes of all strains indicates that the pigment geometry of the LH1 complex in native membranes is unaffected by the presence of an RC and allows us to draw a model representation of the WT, SK Delta LM and SPUHK1 PSU complexes.