ABSTRACT
The strategic redesign of microbial biosynthetic pathways is a compelling route to access molecules of diverse structure and function in a potentially environmentally sustainable fashion. The promise of this approach hinges on an improved understanding of acyl carrier proteins (ACPs), which serve as central hubs in biosynthetic pathways. These small, flexible proteins mediate the transport of molecular building blocks and intermediates to enzymatic partners that extend and tailor the growing natural products. Past combinatorial biosynthesis efforts have failed due to incompatible ACP-enzyme pairings. Herein, we report the design of chimeric ACPs with features of the actinorhodin polyketide synthase ACP (ACT) and of the Escherichia coli fatty acid synthase (FAS) ACP (AcpP). We evaluate the ability of the chimeric ACPs to interact with the E. coli FAS ketosynthase FabF, which represents an interaction essential to building the carbon backbone of the synthase molecular output. Given that AcpP interacts with FabF but ACT does not, we sought to exchange modular features of ACT with AcpP to confer functionality with FabF. The interactions of chimeric ACPs with FabF were interrogated using sedimentation velocity experiments, surface plasmon resonance analyses, mechanism-based cross-linking assays, and molecular dynamics simulations. Results suggest that the residues guiding AcpP-FabF compatibility and ACT-FabF incompatibility may reside in the loop I, α-helix II region. These findings can inform the development of strategic secondary element swaps that expand the enzyme compatibility of ACPs across systems and therefore represent a critical step toward the strategic engineering of "un-natural" natural products.
Subject(s)
Acyl Carrier Protein/metabolism , Escherichia coli Proteins/metabolism , Fatty Acid Synthases/metabolism , Polyketide Synthases/metabolism , Acyl Carrier Protein/chemistry , Amino Acid Sequence , Chimera/metabolism , Escherichia coli/enzymology , Escherichia coli/metabolism , Escherichia coli Proteins/chemistry , Fatty Acid Synthase, Type II/metabolism , Fatty Acid Synthases/chemistry , Fatty Acids/metabolism , Molecular Dynamics Simulation , Polyketide Synthases/chemistry , Polyketides/metabolism , Surface Plasmon Resonance/methods , Transferases (Other Substituted Phosphate Groups)/metabolismABSTRACT
It has been shown that in hybrid polymer-inorganic photovoltaic devices not all the photogenerated excitons dissociate at the interface immediately, but can instead exist temporarily as bound charge pairs (BCPs). Many of these BCPs do not contribute to the photocurrent, as their long lifetime as a bound species promotes various charge carrier recombination channels. Fast and efficient dissociation of BCPs is therefore considered a key challenge in improving the performance of polymer-inorganic cells. Here we investigate the influence of an inorganic energy cascading Nb2O5 interlayer on the charge carrier recombination channels in poly(3-hexylthiophene-2,5-diyl) (P3HT)-TiO2 and PbSe colloidal quantum dot-TiO2 photovoltaic devices. We demonstrate that the additional Nb2O5 film leads to a suppression of BCP formation at the heterojunction of the P3HT cells and also a reduction in the nongeminate recombination mechanisms in both types of cells. Furthermore, we provide evidence that the reduction in nongeminate recombination in the P3HT-TiO2 devices is due in part to the passivation of deep midgap trap states in the TiO2, which prevents trap-assisted Shockley-Read-Hall recombination. Consequently a significant increase in both the open-circuit voltage and the short-circuit current was achieved, in particular for P3HT-based solar cells, where the power conversion efficiency increased by 39%.
