Biophysical chemistry最新文献

β-lactam antibiotics are the most successful and commonly used antibacterial agents, but the emergence of resistance to these drugs has become a global health threat. The expression of β-lactamase enzymes produced by pathogens, which hydrolyze the amide bond of the β-lactam ring, is the major mechanism for bacterial resistance to β-lactams. In particular, among class A, B, C and D β-lactamases, metallo-β-lactamases (MBLs, class B β-lactamases) are considered crucial contributors to resistance in gram-negative bacteria. To combat β-lactamase-mediated resistance, great efforts have been made to develop β-lactamase inhibitors that restore the activity of β-lactams. Some β-lactamase inhibitors, such as diazabicyclooctanes (DBOs) and boronic acid derivatives, have also been approved by the FDA. Inhibitors used in the clinic can inactivate mostly serine-β-lactamases (SBLs, class A, C, and D β-lactamases) but have not been effective against MBLs until now. In order to develop new inhibitors particularly for MBLs, various attempts have been suggested. Based on structural and mechanical studies of MBL enzymes, several MBL inhibitor candidates, including taniborbactam in phase 3 and xeruborbactam in phase 1, have been introduced in recent years. However, designing potent inhibitors that are effective against all subclasses of MBLs is still extremely challenging. This review summarizes not only the types of β-lactamase and mechanisms by which β-lactam antibiotics are inactivated, but also the research finding on β-lactamase inhibitors targeting these enzymes. These detailed information on β-lactamases and their inhibitors could give valuable information for novel β-lactamase inhibitors design.

Nucleic acid aptamers have captivated the attention of analytical and medicinal scientists globally due to their several advantages as recognition molecules over conventional antibodies because of their small size, simple and inexpensive synthesis, broad target range, and high stability in varied environmental conditions. These recognition molecules can be chemically modified to make them resistant to nuclease action in blood serum, reduce rapid renel clearance, improve the target affinity and selectivity, and make them amenable to chemically conjugate with a support system that facilitates their selective applications. This review focuses on the development of efficient aptamer candidates and their application in clinical diagnosis and therapeutic applications. Significant advances have been made in aptamer-based diagnosis of infectious and non-infectious diseases. Collaterally, the progress made in therapeutic applications of aptamers is encouraging, as evident from their use in diagnosing cancer, neurodegenerative diseases, microbial infection, and in imaging. This review also updates the progress on clinical trials of many aptamer-based products of commercial interests. The key development and critical issues on the subject have been summarized in the concluding remarks.

Hydrogenases are a diverse group of metalloenzymes that catalyze the conversion of H₂ into protons and electrons and the reverse reaction. A subgroup is formed by the [FeFe]‑hydrogenases, which are the most efficient enzymes of microbes for catalytic H₂ conversion. We have determined the stability and activity of two [FeFe]‑hydrogenases under high temperature and pressure conditions employing FTIR spectroscopy and the high-pressure stopped-flow methodology in combination with fast UV/Vis detection. Our data show high temperature stability and an increase in activity up to the unfolding temperatures of the enzymes. Remarkably, both enzymes reveal a very high pressure stability of their structure, even up to pressures of several kbars. Their high pressure-stability enables high enzymatic activity up to 2 kbar, which largely exceeds the pressure limit encountered by organisms in the deep sea and sub-seafloor on Earth.

In the realm of biomedical engineering and materials science, the synthesis of biomaterials plays a pivotal role in advancing therapeutic strategies for regeneration of tissues. The deliberate control of crystallization processes in biomaterial synthesis has emerged as a key avenue for tailoring the properties of these materials, enabling the design of innovative solutions for a wide array of medical applications. This review delves into the interplay between controlled crystallization and biomaterial synthesis, exploring its multifaceted applications in the therapeutic domains. The investigation encompasses a wide spectrum of matrices, ranging from small molecules to large biomolecules, highlighting their unique contributions in modulating crystallization processes. Furthermore, the review critically assesses the analytical techniques and methodologies employed to probe and characterize the depths of crystallization dynamics. Advanced imaging, spectroscopic, and computational tools are discussed in the context of unraveling the intricate mechanisms governing nucleation and crystallization processes within the organic matrix. Finally we delve in the applications of such advance material in therapeutics of hard and soft tissues.

In the recent past, there has been an ever-increasing interest in the search for metal-based therapeutic drug candidates for protein misfolding disorders (PMDs) particularly neurodegenerative disorders such as Alzheimer's, Parkinson's, Prion's diseases, and amyotrophic lateral sclerosis. Also, different amyloidogenic variants of human lysozyme (HL) are involved in hereditary systemic amyloidosis. Metallo-therapeutic agents are extensively studied as antitumor agents, however, they are relatively unexplored for the treatment of non-neuropathic amyloidoses. In this work, inhibition potential of a novel ionic cobalt(II) therapeutic agent (CoTA) of the formulation [Co(phen)(H₂O)₄]⁺[glycinate]⁻ is evaluated against HL fibrillation. Various biophysical techniques viz., dye-binding assays, dynamic light scattering (DLS), differential scanning calorimetry (DSC), electron microscopy, and molecular docking experiments validate the proposed mechanism of inhibition of HL fibrillation by CoTA. The experimental corroborative results of these studies reveal that CoTA can suppress and slow down HL fibrillation at physiological temperature and pH. DLS and 1-anilino-8-naphthalenesulfonate (ANS) assay show that reduced fibrillation in the presence of CoTA is marked by a significant decrease in the size and hydrophobicity of the aggregates. Fluorescence quenching and molecular docking results demonstrate that CoTA binds moderately to the aggregation-prone region of HL (K_b = 6.6 × 10⁴ M⁻¹), thereby, inhibiting HL fibrillation. In addition, far-UV CD and DSC show that binding of CoTA to HL does not cause any change in the stability of HL. More importantly, CoTA attenuates membrane damaging effects of HL aggregates against RBCs. This study identifies inorganic metal complexes as a therapeutic intervention for systemic amyloidosis.

Phenylketonuria is characterized by the accumulation of phenylalanine, resulting in severe cognitive and neurological disorders if not treated by a remarkably strict diet. There are two approved drugs today, yet both provide only a partial solution. We have previously demonstrated the formation of amyloid-like toxic assemblies by aggregation of phenylalanine, suggesting a new therapeutic target to be further pursued. Moreover, we showed that compounds that halt the formation of these assemblies also prevent their resulting toxicity. Here, we performed high-throughput screening, searching for compounds with inhibitory effects on phenylalanine aggregation. Morin hydrate, one of the most promising hits revealed during the screen, was chosen to be tested in vivo using a phenylketonuria mouse model. Morin hydrate significantly improved cognitive and motor function with a reduction in the number of phenylalanine brain deposits. Moreover, while phenylalanine levels remained high, we observed a recovery in dopaminergic, adrenergic, and neuronal markers. To conclude, the ability of Morin hydrate to halt phenylalanine aggregation without reducing phenylalanine levels implies the toxic role of the phenylalanine assemblies in phenylketonuria and opens new avenues for disease-modifying treatment.

Micro- and nanoplastics have become a significant concern, due to their ubiquitous presence in the environment. These particles can be internalized by the human body through ingestion, inhalation, or dermal contact, and then they can interact with environmental or biological molecules, such as proteins, resulting in the formation of the protein corona. However, information on the role of protein corona in the human body is still missing. Coarse-grain models of the nanoplastics and pentapeptides were created and simulated at the microscale to study the role of protein corona. Additionally, a lipid bilayer coarse-grain model was reproduced to investigate the behavior of the coronated nanoplastics in proximity of a lipid bilayer. Hydrophobic and aromatic amino acids have a high tendency to create stable bonds with all nanoplastics. Moreover, polystyrene and polypropylene establish bonds with polar and charged amino acids. When the coronated nanoplastics are close to a lipid bilayer, different behaviors can be observed. Polyethylene creates a single polymeric chain, while polypropylene tends to break down into its single chains. Polystyrene can both separate into its individual chains and remain aggregated. The protein corona plays an important role when interacting with the nanoplastics and the lipid membrane. More studies are needed to validate the results and to enhance the complexity of the systems.

Boundary lipids surrounding membrane proteins play an essential role in protein function and structure. These protein–lipid interactions are mainly divided into electrostatic interactions between the polar amino acids of proteins and polar heads of phospholipids, and hydrophobic interactions between protein transmembrane sites and phospholipid acyl chains. Our previous report (Kawatake et al., Biochim. Biophys. Acta 1858 [2016] 2106–2115) covered a method for selectively analyzing boundary lipid interactions and showed differences in membrane protein–peripheral lipid interactions due to differences in their head group. Interactions in the hydrophobic acyl chains of phospholipids are relatively consistent among proteins, but the details of these interactions have not been elucidated. In this study, we reconstituted bacteriorhodopsin as a model protein into phospholipid membranes labeled with ²H and ¹³C for solid-state NMR measurement to investigate the depth-dependent effect of the head group structure on the lipid bilayer. The results showed that the position of the phospholipid near the carbonyl carbon was affected by the head group in terms of selectivity for protein surfaces, whereas in the deep interior of the bilayer near the leaflet interface, there was little difference between the head groups, indicating that the dependence of their interactions on the head group was much reduced.

Spin-labeling with electron paramagnetic resonance spectroscopy (EPR) is a facile method for interrogating macromolecular flexibility, conformational changes, accessibility, and hydration. Within we present a computationally based approach for the rational selection of reporter sites in Bacillus subtilis lipase A (BSLA) for substitution to cysteine residues with subsequent modification with a spin-label that are expected to not significantly perturb the wild-type structure, dynamics, or enzymatic function. Experimental circular dichroism spectroscopy, Michaelis-Menten kinetic parameters and EPR spectroscopy data validate the success of this approach to computationally select reporter sites for future magnetic resonance investigations of hydration and hydration changes induced by polymer conjugation, tethering, immobilization, or amino acid substitution in BSLA. Analysis of molecular dynamic simulations of the impact of substitutions on the secondary structure agree well with experimental findings. We propose that this computationally guided approach for choosing spin-labeled EPR reporter sites, which evaluates relative surface accessibility coupled with hydrogen bonding occupancy of amino acids to the catalytic pocket via atomistic simulations, should be readily transferable to other macromolecular systems of interest including selecting sites for paramagnetic relaxation enhancement NMR studies, other spin-labeling EPR studies or any method requiring a tagging method where it is desirable to not alter enzyme stability or activity.

Amyloid and amorphous aggregates represent the two major categories of aggregates associated with diseases, and although exhibiting distinct features, researchers often treat them as equivalent, which demonstrates the need for more thorough characterization. Here, we compare amyloid and amorphous aggregates based on their biochemical properties, kinetics, and morphological features. To further decipher this issue, we propose the use of peptide self-assemblies as minimalistic models for understanding the aggregation process. Peptide building blocks are significantly smaller than proteins that participate in aggregation, however, they make a plausible means to bridge the gap in discerning the aggregation process at the more complex, protein level. Additionally, we explore the potential use of peptide-inspired models to research the liquid-liquid phase separation as a feasible mechanism preceding amyloid formation. Connecting these concepts can help clarify our understanding of aggregation-related disorders and potentially provide novel drug targets to impede and reverse these serious illnesses.