Two pathways to understanding electron transfer in reaction centers from photosynthetic bacteria: A comparison of Rhodobacter sphaeroides and Rhodobacter capsulatus mutants.

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+HB- yields (up to ∼90%) than their R. capsulatus analogs (up to ∼60%), where HB is the B-side bacteriopheophytin. Substitution of Tyr for Phe at L-polypeptide position L181 near BB primarily increases the contribution of fast P* â†’ P+BB- â†’ P+HB- two-step ET, where BB 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* â†’ P+BA- â†’ P+HA-) where the second step (∼1 ps) is faster than the first (∼3-4 ps) in the native RC. Substitutions near HB, 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* â†’ P+HB- one-step superexchange ET. Both ET mechanisms are effective in directing electrons "the wrong way" to HB 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.

High Yield of B-Side Electron Transfer at 77 K in the Photosynthetic Reaction Center Protein from Rhodobacter sphaeroides.

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 C2-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+BB- → P+HB-), one-step superexchange ET (e.g., P* → P+HB-), and P* → ground state. [HB and BB 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+HB- 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+HB-, 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.

A reinforcement learning framework for pooled oligonucleotide design.

MOTIVATION: The goal of oligonucleotide (oligo) design is to select oligos that optimize a set of design criteria. Oligo design problems are combinatorial in nature and require computationally intensive models to evaluate design criteria. Even relatively small problems can be intractable for brute-force approaches that test every possible combination of oligos, so heuristic approaches must be used to find near-optimal solutions. RESULTS: We present a general reinforcement learning (RL) framework, called OligoRL, to solve oligo design problems with complex constraints. OligoRL allows 'black-box' design criteria and can be adapted to solve many oligo design problems. We highlight the flexibility of OligoRL by building tools to solve three distinct design problems: (i) finding pools of random DNA barcodes that lack restriction enzyme recognition sequences (CutFreeRL); (ii) compressing large, non-degenerate oligo pools into smaller degenerate ones (OligoCompressor) and (iii) finding Not-So-Random hexamer primer pools that avoid rRNA and other unwanted transcripts during RNA-seq library preparation (NSR-RL). OligoRL demonstrates how RL offers a general solution for complex oligo design problems. AVAILABILITY AND IMPLEMENTATION: OligoRL and all simulation codes are available as a Julia package at http://jensenlab.net/tools and archived at https://archive.softwareheritage.org/browse/origin/directory/?origin_url=https://github.com/bmdavid2/OligoRL. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Species differences in unlocking B-side electron transfer in bacterial reaction centers.

The structure of the bacterial photosynthetic reaction center (RC) reveals symmetry-related electron transfer (ET) pathways, but only one path is used in native RCs. Analogous mutations have been made in two Rhodobacter (R.) species. A glutamic acid at position 133 in the M subunit increases transmembrane charge separation via the naturally inactive (B-side) path through impacts on primary ET in mutant R. sphaeroidesRCs. Prior work showed that the analogous substitution in the R. capsulatusRC also increases B-side activity, but mainly affects secondary ET. The overall yields of transmembrane ET are similar, but enabled in fundamentally different ways.