Search

Whereas modern proteins rely on a quasi-universal repertoire of 20 canonical amino acids (AAs), numerous lines of evidence suggest that ancient proteins relied on a limited alphabet of 10 ‘early’ AAs, and that the 10 ‘late’ AAs were products of biosynthetic pathways. However, many non-proteinogenic AAs were also prebiotically available, which begs two fundamental questions: Why do we have the current modern amino acid alphabet, and Would proteins be able to fold into globular structures as well if different amino acids comprised the genetic code? Here, we experimentally evaluated the solubility and secondary structure propensities of several prebiotically relevant amino acids in the context of synthetic combinatorial 25-mer peptide libraries. The most prebiotically abundant linear aliphatic and basic residues were incorporated along with or in place of other early amino acids to explore these alternative sequence spaces. We show that foldability was a critical factor in the selection of the canonical alphabet. Unbranched aliphatic and short-chain basic amino acids were purged from the proteinogenic alphabet despite their high prebiotic abundance because they generate polypeptides that are over-solubilized and have low packing efficiency. Surprisingly, we find that the inclusion of a short-chain basic amino acid also decreases polypeptides’ secondary structure potential. Our results support the view that despite lacking basic residues, the early canonical alphabet was remarkably adaptive at supporting protein folding and explain why basic residues were only incorporated at a later stage of the alphabet evolution.

Evaluating the significance of various forms of DNA damage is complicated by discoveries that some lesions inactivate repair enzymes or produce more deleterious forms of damage. Histone lysines within nucleosomes react with the commonly produced C4′-oxidized abasic site (C4-AP) to concomitantly yield an electrophilic modification (K(MP)) on lysine and DNA strand scission. We developed a chemoproteomic approach to identify K(MP) in HeLa cells. More than 60 000 K(MP)-modified histones are produced per cell. Using LC-MS/MS, we detected K(MP) at 17 of the 57 lysine residues distributed throughout the four core histone proteins. Therefore, K(MP) constitutes a DNA damage-induced, nonenzymatic histone post-translational modification. K(MP) formation suggests that downstream processes resulting from DNA damage could have ramifications on cells.

Decades of research on protein folding have primarily focused on a subset of small proteins that can reversibly refold from a denatured state. However, these studies have generally not been representative of the complexity of natural proteomes, which consist of many proteins with complex architectures and domain organizations. Here, we introduce an experimental approach to probe protein refolding kinetics for whole proteomes using mass spectrometry-based proteomics. Our study covers the majority of the soluble E. coli proteome expressed during log-phase growth, and among this group, we find that one-third of the E. coli proteome is not intrinsically refoldable on physiological time scales, a cohort that is enriched with certain fold-types, domain organizations, and other biophysical features. We also identify several properties and fold-types that are correlated with slow refolding on the minute time scale. Hence, these results illuminate when exogenous factors and processes, such as chaperones or cotranslational folding, might be required for efficient protein folding.

Electrostatic interactions play a pivotal role in enzymatic catalysis and are increasingly modeled explicitly in computational enzyme design; nevertheless, they are challenging to measure experimentally. Using vibrational Stark effect (VSE) spectroscopy, we have measured electric fields inside the active site of the enzyme ketosteroid isomerase (KSI). These studies have shown that these fields can be unusually large, but it has been unclear to what extent they specifically stabilize the transition state (TS) relative to a ground state (GS). In the following, we use crystallography and computational modeling to show that KSI’s intrinsic electric field is nearly perfectly oriented to stabilize the geometry of its reaction’s TS. Moreover, we find that this electric field adjusts the orientation of its substrate in the ground state so that the substrate needs to only undergo minimal structural changes upon activation to its TS. This work provides evidence that the active site electric field in KSI is preorganized to facilitate catalysis and provides a template for how electrostatic preorganization can be measured in enzymatic systems.

Synthetic methods that edit soft polymer backbones are critical technologies for tailoring the structures and properties of macromolecules. Developing strategies that leverage underexplored reaction manifolds are vital for accessing new chemical (and functional) space in soft materials. Here, we report a mild electrochemical approach that enables both degradation and functionalization of synthetic polymers. We found that bulk electrolysis (under either homogeneous or heterogeneous conditions) promoted facile, chemoselective chain scission in a variety of olefin-containing materials. Polymer degradation could also be coupled with functionalization (e.g., azidation) to afford new species that could serve as macromonomers.

Decades of research on protein folding have primarily focused on a subset of small proteins that can reversibly refold from a denatured state. However, these studies have generally not been representative of the complexity of natural proteomes, which consist of many proteins with complex architectures and domain organizations. Here, we introduce an experimental approach to probe protein refolding kinetics for whole proteomes using mass spectrometry-based proteomics. Our study covers the majority of the soluble E. coli proteome expressed during log-phase growth, and among this group, we find that one third of the E. coli proteome is not intrinsically refoldable on physiological timescales, a cohort that is enriched with certain fold-types, domain organizations, and other biophysical features. We also identify several properties and fold-types that correlate with slow refolding on the minute timescale. Hence, these results illuminate when exogenous factors and processes, such as chaperones or co-translational folding, might be required for efficient protein folding.

Vibrational probes can provide a direct read-out of the local electrostatic field in complex molecular environments, such as protein binding sites and enzyme active sites. This information provides an experimental method to explore the underlying physical causes of important biomolecular processes such as binding and catalysis. However, specific chemical interactions such as hydrogen bonds can have complicated effects on vibrational probes and confound simple electrostatic interpretations of their frequency shifts. We employ vibrational Stark spectroscopy along with infrared spectroscopy of carbonyl probes in different solvent environments and in Ribonuclease S to understand the sensitivity of carbonyl frequencies to electrostatic fields, including those due to hydrogen bonds. Additionally, we carried out molecular dynamics simulations to calculate ensemble-averaged electric fields in solvents and in Ribonuclease S, and found excellent correlation between calculated fields and vibrational frequencies. These data enabled the construction of a robust field-frequency calibration curve for the C=O vibration. The present results suggest that carbonyl probes are capable of quantitatively assessing the electrostatics of hydrogen bonding, making them promising for future study of protein function.

Electrostatic interactions provide a primary connection between a protein's three-dimensional structure and its function. Infrared probes are useful because vibrational frequencies of certain chemical groups, such as nitriles, are linearly sensitive to local electrostatic field and can serve as a molecular electric field meter. IR spectroscopy has been used to study electrostatic changes or fluctuations in proteins, but measured peak frequencies have not been previously mapped to total electric fields, because of the absence of a field-frequency calibration and the complication of local chemical effects such as H-bonds. We report a solvatochromic model that provides a means to assess the H-bonding status of aromatic nitrile vibrational probes and calibrates their vibrational frequencies to electrostatic field. The analysis involves correlations between the nitrile's IR frequency and its (13)C chemical shift, whose observation is facilitated by a robust method for introducing isotopes into aromatic nitriles. The method is tested on the model protein ribonuclease S (RNase S) containing a labeled p-CN-Phe near the active site. Comparison of the measurements in RNase S against solvatochromic data gives an estimate of the average total electrostatic field at this location. The value determined agrees quantitatively with molecular dynamics simulations, suggesting broader potential for the use of IR probes in the study of protein electrostatics.

We report the first quantum-based multiscale simulations to study the reactivity of shocked perfect crystals of the insensitive energetic material triaminotrinitrobenzene (TATB). Tracking chemical transformations of TATB experiencing overdriven shock speeds of 9 km/s for up to 0.43 ns and 10 km/s for up to 0.2 ns reveal high concentrations of nitrogen-rich heterocyclic clusters. Further reactivity of TATB toward the final decomposition products of fluid N(2) and solid carbon is inhibited due to the formation of these heterocycles. Our results thus suggest a new mechanism for carbon-rich explosive materials that precedes the slow diffusion-limited process of forming the bulk solid from carbon clusters and provide fundamental insight at the atomistic level into the long reaction zone of shocked TATB.

Magnesium-catalyzed enantioselective aldol between ethyl diazoacetate and aromatic, aliphatic, and alpha,beta-unsaturated aldehydes affords alpha-diazo-beta-hydroxy-esters in high enantioselectivities. Aldol adducts resulting from this asymmetric transformation are versatile intermediates toward the synthesis of several ester containing chiral building blocks.

Synthese a partir de la reaction d'(alcynyl trialkylsilyl) ethers avec le ([difluoro-4,4 iodo-4 butyne-2 yl] [perhydro pyrannyl-2]) ether en presence de butyl-lithium

Report discussing methods for preparing various protactinium compounds, including protactinium oxides, tetrafluoride, and oxysulfide. Additionally, a method for making the protactinium metal is mentioned.

2 methods are described for the preparation of an oxygen functionalized vinylcopper reagent. Reactions of this reagent with cyclic and acyclic enones give products of 14 addition. The labile methoxyisopropyl group was used as an alcohol protecting group for ease of formation and removal. The influence of reaction conditions such as solvents and temperature on the mode of addition and yield is discussed. (S)-1-Iodo-trans-1-octen-3-ol (16a) was prepared from (S)-1-octyn-3-ol (17). The optically pure iodovinylcarbinol was converted to the cuprate 2 and 14 addition to the hydroxy-protected cyclopentenone 14c afforded (-)-PGE1 (18b). (Authors modified)