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1. Intrinsic electronic conductivity of individual atomically resolved amyloid crystals reveals micrometer-long hole hopping via tyrosines

   https://doi.org/10.1073/pnas.2014139118  

   Shipps, Catharine; Kelly, H. Ray; Dahl, Peter J.; Yi, Sophia M.; Vu, Dennis; Boyer, David; Glynn, Calina; Sawaya, Michael R.; Eisenberg, David; Batista, Victor S.; et al (December 2020, Proceedings of the National Academy of Sciences)

   Proteins are commonly known to transfer electrons over distances limited to a few nanometers. However, many biological processes require electron transport over far longer distances. For example, soil and sediment bacteria transport electrons, over hundreds of micrometers to even centimeters, via putative filamentous proteins rich in aromatic residues. However, measurements of true protein conductivity have been hampered by artifacts due to large contact resistances between proteins and electrodes. Using individual amyloid protein crystals with atomic-resolution structures as a model system, we perform contact-free measurements of intrinsic electronic conductivity using a four-electrode approach. We find hole transport through micrometer-long stacked tyrosines at physiologically relevant potentials. Notably, the transport rate through tyrosines (10⁵s⁻¹) is comparable to cytochromes. Our studies therefore show that amyloid proteins can efficiently transport charges, under ordinary thermal conditions, without any need for redox-active metal cofactors, large driving force, or photosensitizers to generate a high oxidation state for charge injection. By measuring conductivity as a function of molecular length, voltage, and temperature, while eliminating the dominant contribution of contact resistances, we show that a multistep hopping mechanism (composed of multiple tunneling steps), not single-step tunneling, explains the measured conductivity. Combined experimental and computational studies reveal that proton-coupled electron transfer confers conductivity; both the energetics of the proton acceptor, a neighboring glutamine, and its proximity to tyrosine influence the hole transport rate through a proton rocking mechanism. Surprisingly, conductivity increases 200-fold upon cooling due to higher availability of the proton acceptor by increased hydrogen bonding. 

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2. A promiscuous cytochrome P450 aromatic O-demethylase for lignin bioconversion

   https://doi.org/10.1038/s41467-018-04878-2  

   Mallinson, Sam J. B.; Machovina, Melodie M.; Silveira, Rodrigo L.; Garcia-Borràs, Marc; Gallup, Nathan; Johnson, Christopher W.; Allen, Mark D.; Skaf, Munir S.; Crowley, Michael F.; Neidle, Ellen L.; et al (June 2018, Nature Communications)

   Abstract Microbial aromatic catabolism offers a promising approach to convert lignin, a vast source of renewable carbon, into useful products. Aryl-O-demethylation is an essential biochemical reaction to ultimately catabolize coniferyl and sinapyl lignin-derived aromatic compounds, and is often a key bottleneck for both native and engineered bioconversion pathways. Here, we report the comprehensive characterization of a promiscuous P450 aryl-O-demethylase, consisting of a cytochrome P450 protein from the family CYP255A (GcoA) and a three-domain reductase (GcoB) that together represent a new two-component P450 class. Though originally described as converting guaiacol to catechol, we show that this system efficiently demethylates both guaiacol and an unexpectedly wide variety of lignin-relevant monomers. Structural, biochemical, and computational studies of this novel two-component system elucidate the mechanism of its broad substrate specificity, presenting it as a new tool for a critical step in biological lignin conversion. 

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3. pLMSNOSite: an ensemble-based approach for predicting protein S-nitrosylation sites by integrating supervised word embedding and embedding from pre-trained protein language model

   Pawel, Pratyush; Suresh, Pokharel; Hiroto, Saigo; Dukka, KC (February 2023, BMC bioinformatics)

   Protein S-nitrosylation (SNO) plays a key role in transferring nitric oxide-mediated signals in both animals and plants and has emerged as an important mechanism for regulating protein functions and cell signaling of all main classes of protein. It is involved in several biological processes including immune response, protein stability, transcription regulation, post translational regulation, DNA damage repair, redox regulation, and is an emerging paradigm of redox signaling for protection against oxidative stress. The development of robust computational tools to predict protein SNO sites would contribute to further interpretation of the pathological and physiological mechanisms of SNO. 

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4. Enabling microbial syringol conversion through structure-guided protein engineering

   https://doi.org/10.1073/pnas.1820001116  

   Machovina, Melodie M.; Mallinson, Sam J.; Knott, Brandon C.; Meyers, Alexander W.; Garcia-Borràs, Marc; Bu, Lintao; Gado, Japheth E.; Oliver, April; Schmidt, Graham P.; Hinchen, Daniel J.; et al (January 2019, Proceedings of the National Academy of Sciences)

   Microbial conversion of aromatic compounds is an emerging and promising strategy for valorization of the plant biopolymer lignin. A critical and often rate-limiting reaction in aromatic catabolism is O -aryl-demethylation of the abundant aromatic methoxy groups in lignin to form diols, which enables subsequent oxidative aromatic ring-opening. Recently, a cytochrome P450 system, GcoAB, was discovered to demethylate guaiacol (2-methoxyphenol), which can be produced from coniferyl alcohol-derived lignin, to form catechol. However, native GcoAB has minimal ability to demethylate syringol (2,6-dimethoxyphenol), the analogous compound that can be produced from sinapyl alcohol-derived lignin. Despite the abundance of sinapyl alcohol-based lignin in plants, no pathway for syringol catabolism has been reported to date. Here we used structure-guided protein engineering to enable microbial syringol utilization with GcoAB. Specifically, a phenylalanine residue (GcoA-F169) interferes with the binding of syringol in the active site, and on mutation to smaller amino acids, efficient syringol O -demethylation is achieved. Crystallography indicates that syringol adopts a productive binding pose in the variant, which molecular dynamics simulations trace to the elimination of steric clash between the highly flexible side chain of GcoA-F169 and the additional methoxy group of syringol. Finally, we demonstrate in vivo syringol turnover in Pseudomonas putida KT2440 with the GcoA-F169A variant. Taken together, our findings highlight the significant potential and plasticity of cytochrome P450 aromatic O -demethylases in the biological conversion of lignin-derived aromatic compounds. 

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5. Breakdown of the Small‐Polaron Hopping Model in Higher‐Order Spinels

   https://doi.org/10.1002/adma.202004490  

   Bhargava, Anuj; Eppstein, Roni; Sun, Jiaxin; Smeaton, Michelle_A; Paik, Hanjong; Kourkoutis, Lena_F; Schlom, Darrell_G; Caspary_Toroker, Maytal; Robinson, Richard_D (October 2020, Advanced Materials)

   Abstract The small‐polaron hopping model has been used for six decades to rationalize electronic charge transport in oxides. The model was developed for binary oxides, and, despite its significance, its accuracy has not been rigorously tested for higher‐order oxides. Here, the small‐polaron transport model is tested by using a spinel system with mixed cation oxidation states (Mn_xFe_3−xO₄). Using molecular‐beam epitaxy (MBE), a series of single crystal Mn_xFe_3−xO₄thin films with controlled stoichiometry, 0 ≤x ≤ 2.3, and lattice strain are grown, and the cation site‐occupation is determined through X‐ray emission spectroscopy (XES). Density functional theory +Uanalysis shows that charge transport occurs only between like‐cations (Fe/Fe or Mn/Mn). The site‐occupation data and percolation models show that there are limited stoichiometric ranges for transport along Fe and Mn pathways. Furthermore, due to asymmetric hopping barriers and formation energies, the polaron is energetically preferred to the polaron, resulting in an asymmetric contribution of Mn/Mn pathways. All of these findings are not contained in the conventional small‐polaron hopping model, highlighting its inadequacy. To correct the model, new parameters in the nearest‐neighbor hopping equation are introduced to account for percolation, cross‐hopping, and polaron‐distribution, and it is found that a near‐perfect correlation can be made between experiment and theory for the electronic conductivity. 

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