PBP-A, a cyanobacterial DD-peptidase with high specificity for amidated muropeptides, exhibits pH-dependent promiscuous activity harmful to Escherichia coli.

Penicillin binding proteins (PBPs) are involved in biosynthesis, remodeling and recycling of peptidoglycan (PG) in bacteria. PBP-A from Thermosynechococcus elongatus belongs to a cyanobacterial family of enzymes sharing close structural and phylogenetic proximity to class A ß-lactamases. With the long-term aim of converting PBP-A into a ß-lactamase by directed evolution, we simulated what may happen when an organism like Escherichia coli acquires such a new PBP and observed growth defect associated with the enzyme activity. To further explore the molecular origins of this harmful effect, we decided to characterize deeper the activity of PBP-A both in vitro and in vivo. We found that PBP-A is an enzyme endowed with DD-carboxypeptidase and DD-endopeptidase activities, featuring high specificity towards muropeptides amidated on the D-iso-glutamyl residue. We also show that a low promiscuous activity on non-amidated peptidoglycan deteriorates E. coli's envelope, which is much higher under acidic conditions where substrate discrimination is mitigated. Besides expanding our knowledge of the biochemical activity of PBP-A, this work also highlights that promiscuity may depend on environmental conditions and how it may hinder rather than promote enzyme evolution in nature or in the laboratory.

Impact of Deamidation on the Structure and Function of Antiapoptotic Bcl-xL.

Bcl-xL is an antiapoptotic mitochondrial trans-membrane protein, which is known to play a crucial role in the survival of tumor cells. The deamidation of Bcl-xL is a pivotal switch that regulates its biological function. The potential impact of deamidation on the structure and dynamics of Bcl-xL is directly linked to the intrinsically disordered region (IDR), which is the main site for post-translational modifications (PTMs). In this study, we explored deamidation-induced conformational changes in Bcl-xL to gain insight into its loss of function by performing microsecond-long molecular dynamics (MD) simulations. MD simulation outcomes showed that the IDR motion and interaction patterns have changed notably upon deamidation. Principal component analysis (PCA) demonstrates significant differences between wild-type and deamidated Bcl-xL and suggests that deamidation affects the structure and dynamics of Bcl-xL. The combination of clustering analysis, H-bond analysis, and PCA revealed changes in conformation, interaction, and dynamics upon deamidation. Differences in contact patterns and essential dynamics that lead to a narrowing in the binding groove (BG) are clear indications of deamidation-induced allosteric effects. In line with previous studies, we show that the IDR plays a very important role in the loss of apoptotic functions of Bcl-xL while providing a unique perspective on the underlying mechanism of Bcl-xL deamidation-induced cell death.

Keteniminium Salts: Reactivity and Propensity toward Electrocyclization Reactions.

A predictive computational study was conducted in order to assess the efficiency of electrocyclization reactions of keteniminium salts, in an effort to form a variety of heterocyclic systems, namely, 3-amino(benzo)thiophenes, 3-amino(benzo)furans, 3-aminopyrroles, as well as 3-aminoindoles. A density functional theory (DFT) approach was utilized and the effect of heteroatoms (NMe, O, S) was thoroughly investigated by means of population analysis, QTAIM, NICS, ACID, and local reactivity descriptors (Parr and Fukui functions). The electrocyclization of enamines leading to 3-aminopyrroles was shown to be both kinetically and thermodynamically most favorable. Moreover, the pericyclic nature of the electrocyclizations was confirmed using FMO, QTAIM, NICS, and ACID methods. Additionally, substituent effects were investigated in order to give further insight on the reactivity of heteroatom containing keteniminium systems toward electrocyclization reactions. Finally, computational predictions were experimentally confirmed for a selection of keteniminium systems.

Theoretical insight into the regioselective ring-expansions of bicyclic aziridinium ions.

Transient bicyclic aziridinium ions are known to undergo ring-expansion reactions, paving the way to functionalized nitrogen-containing heterocycles. In this study, the regioselectivity observed in the ring-expansion reactions of 1-azoniabicyclo[n.1.0]alkanes was investigated from a computational viewpoint to study the ring-expansion pathways of two bicyclic systems with different ring sizes. Moreover, several nucleophiles leading to different experimental results were investigated. The effect of solvation was taken into account using both explicit and implicit solvent models. This theoretical rationalization provides valuable insight into the observed regioselectivity and may be used as a predictive tool in future studies.

Computational Studies on Cinchona Alkaloid-Catalyzed Asymmetric Organic Reactions.

Remarkable progress in the area of asymmetric organocatalysis has been achieved in the last decades. Cinchona alkaloids and their derivatives have emerged as powerful organocatalysts owing to their reactivities leading to high enantioselectivities. The widespread usage of cinchona alkaloids has been attributed to their nontoxicity, ease of use, stability, cost effectiveness, recyclability, and practical utilization in industry. The presence of tunable functional groups enables cinchona alkaloids to catalyze a broad range of reactions. Excellent experimental studies have extensively contributed to this field, and highly selective reactions were catalyzed by cinchona alkaloids and their derivatives. Computational modeling has helped elucidate the mechanistic aspects of cinchona alkaloid catalyzed reactions as well as the origins of the selectivity they induce. These studies have complemented experimental work for the design of more efficient catalysts. This Account presents recent computational studies on cinchona alkaloid catalyzed organic reactions and the theoretical rationalizations behind their effectiveness and ability to induce selectivity. Valuable efforts to investigate the mechanisms of reactions catalyzed by cinchona alkaloids and the key aspects of the catalytic activity of cinchona alkaloids in reactions ranging from pharmaceutical to industrial applications are summarized. Quantum mechanics, particularly density functional theory (DFT), and molecular mechanics, including ONIOM, were used to rationalize experimental findings by providing mechanistic insights into reaction mechanisms. B3LYP with modest basis sets has been used in most of the studies; nonetheless, the energetics have been corrected with higher basis sets as well as functionals parametrized to include dispersion M05-2X, M06-2X, and M06-L and functionals with dispersion corrections. Since cinchona alkaloids catalyze reactions by forming complexes with substrates via hydrogen bonds and long-range interactions, the use of split valence triple-ζ basis sets including diffuse and polarization functions on heavy atoms and polarization functions on hydrogens are recommended. Most of the studies have used the continuum-based models to mimic the condensed phase in which organocatalysts function; in some cases, explicit solvation was shown to yield better quantitative agreement with experimental findings. The conformational behavior of cinchona alkaloids is also highlighted as it is expected to shed light on the origin of selectivity and pave the way to a comprehensive understanding of the catalytic mechanism. The ultimate goal of this Account is to provide an up-to-date overlook on cinchona alkaloid catalyzed chemistry and provide insight for future studies in both experimental and theoretical fields.