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The rates, yields, mechanisms and directionality of electron transfer (ET) are explored in twelve pairs of Rhodobacter (R.) sphaeroides and R. capsulatus mutant RCs designed to defeat ET from the excited primary donor (P*) to the A-side cofactors and re-direct ET to the normally inactive mirror-image B-side cofactors. In general, the R. sphaeroides variants have larger P + H B - yields (up to ∼90%) than their R. capsulatus analogs (up to ∼60%), where H B is the B-side bacteriopheophytin. Substitution of Tyr for Phe at L-polypeptide position L181 near B B primarily increases the contribution of fast P* → P + B B -  → P + H B - two-step ET, where B B is the "bridging" B-side bacteriochlorophyll. The second step (∼6-8 ps) is slower than the first (∼3-4 ps), unlike A-side two-step ET (P* → P + B A -  → P + H A - ) where the second step (∼1 ps) is faster than the first (∼3-4 ps) in the native RC. Substitutions near H B , at L185 (Leu, Trp or Arg) and at M-polypeptide site M133/131 (Thr, Val or Glu), strongly affect the contribution of slower (20-50 ps) P* → P + H B - one-step superexchange ET. Both ET mechanisms are effective in directing electrons "the wrong way" to H B and both compete with internal conversion of P* to the ground state (∼200 ps) and ET to the A-side cofactors. Collectively, the work demonstrates cooperative amino-acid control of rates, yields and mechanisms of ET in bacterial RCs and how A- vs. B-side charge separation can be tuned in both species., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The primary electron transfer (ET) processes at 295 and 77 K are compared for the Rhodobacter sphaeroides reaction center (RC) pigment-protein complex from 13 mutants including a wild-type control. The engineered RCs bear mutations in the L and M polypeptides that largely inhibit ET from the excited state P* of the primary electron donor (P, a bacteriochlorophyll dimer) to the normally photoactive A-side cofactors and enhance ET to the C 2 -symmetry related, and normally photoinactive, B-side cofactors. P* decay is multiexponential at both temperatures and modeled as arising from subpopulations that differ in contributions of two-step ET ( e.g., P* → P + B B - → P + H B - ), one-step superexchange ET ( e.g., P* → P + H B - ), and P* → ground state. [H B and B B are monomeric bacteriopheophytin and bacteriochlorophyll, respectively.] The relative abundances of the subpopulations and the inherent rate constants of the P* decay routes vary with temperature. Regardless, ET to produce P + H B - is generally faster at 77 K than at 295 K by about a factor of 2. A key finding is that the yield of P + H B - , which ranges from ∼5% to ∼90% among the mutant RCs, is essentially the same at 77 K as at 295 K in each case. Overall, the results show that ET from P* to the B-side cofactors in these mutants does not require thermal activation and involves combinations of ET mechanisms analogous to those operative on the A side in the native RC.

Bacteria belonging to the Lachnospiraceae family are abundant, obligate anaerobic members of the microbiota in healthy humans. Lachnospiraceae impact their hosts by producing short-chain fatty acids, converting primary to secondary bile acids, and facilitating colonization resistance against intestinal pathogens. To increase our understanding of genomic and functional diversity between members of this family, we cultured 273 Lachnospiraceae isolates representing 11 genera and 27 species from human donors and performed whole-genome sequencing assembly and annotation. This analysis revealed substantial inter- and intra-species diversity in pathways that likely influence an isolate's ability to impact host health. These differences are likely to impact colonization resistance through lantibiotic expression or intestinal acidification, influence host mucosal immune cells and enterocytes via butyrate production, or contribute to synergism within a consortium by heterogenous polysaccharide metabolism. Identification of these specific functions could facilitate development of probiotic bacterial consortia that drive and/or restore in vivo microbiome functions., Competing Interests: Declaration of Interests E.G.P. has received speaker honoraria from Bristol-Myer Squibb, Celgene, Seres Therapeutics, MedImmune, Novartis, and Ferring Pharmaceuticals; is an inventor on patent application no. WPO2015179437A1, entitled “Methods and compositions for reducing Clostridium difficile infection” and no. WPO2017091753A1, entitled “Methods and compositions for reducing vancomycin-resistant Enterococci infection or colonization”; and holds patents that receive royalties from Seres Therapeutics. The remaining authors declare no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

In bacterial reaction centers (RCs), photon-induced initial charge separation uses an A-side bacteriochlorophyll (BChl, B A ) and bacteriopheophytin (BPh, H A ), while the near-mirror image B-side B B and H B cofactors are inactive. Two new sets of Rhodobacter capsulatus RC mutants were designed, both bearing substitution of all amino acids for the native histidine M180 (M-polypeptide residue 180) ligand to the core Mg ion of B B . Residues are identified that largely result in retention of a BChl in the B B site (Asp, Ser, Pro, Gln, Asn, Gly, Cys, Lys, and Thr), ones that largely harbor the Mg-free BPh in the B B site (Leu and Ile), and ones for which isolated RCs are comprised of a substantial mixture of these two RC types (Ala, Glu, Val, Met and, in one set, Arg). No protein was isolated when M180 is Trp, Tyr, Phe, or (in one set) Arg. These findings are corroborated by ground state spectra, pigment extractions, ultrafast transient absorption studies, and the yields of B-side transmembrane charge separation. The changes in coordination chemistries did not reveal an RC with sufficiently precise poising of the redox properties of the B B -site cofactor to result in a high yield of B-side electron transfer to H B . Insights are gleaned into the amino acid properties that support BChl in the B B site and into the widely observed multi-exponential decay of the excited state of the primary electron donor. The results also have direct implications for tuning free energies of the charge-separated intermediates in RCs and mimetic systems.