Catalytic and biocatalytic degradation of microplastics.

In recent years, there has been a surge in annual plastic production, which has contributed to growing environmental challenges, particularly in the form of microplastics. Effective management of plastic and microplastic waste has become a critical concern, necessitating innovative strategies to address its impact on ecosystems and human health. In this context, catalytic degradation of microplastics emerges as a pivotal approach that holds significant promise for mitigating the persistent effects of plastic pollution. In this article, we critically explored the current state of catalytic degradation of microplastics and discussed the definition of degradation, characterization methods for degradation products, and the criteria for standard sample preparation. Moreover, the significance and effectiveness of various catalytic entities, including enzymes, transition metal ions (for the Fenton reaction), nanozymes, and microorganisms are summarized. Finally, a few key issues and future perspectives regarding the catalytic degradation of microplastics are proposed.

Oxocarbon Acids and their Derivatives in Biological and Medicinal Chemistry.

The biological and medicinal chemistry of the oxocarbon acids 2,3- dihydroxycycloprop-2-en-1-one (deltic acid), 3,4-dihydroxycyclobut-3-ene-1,2-dione (squaric acid), 4,5-dihydroxy-4-cyclopentene-1,2,3-trione (croconic acid), 5,6-dihydroxycyclohex- 5-ene-1,2,3,4-tetrone (rhodizonic acid) and their derivatives is reviewed and their key chemical properties and reactions are discussed. Applications of these compounds as potential bioisosteres in biological and medicinal chemistry are examined. Reviewed areas include cell imaging, bioconjugation reactions, antiviral, antibacterial, anticancer, enzyme inhibition, and receptor pharmacology.

Molecular Engineering of E. coli Bacterioferritin: A Versatile Nanodimensional Protein Cage.

Currently, intense interest is focused on the discovery and application of new multisubunit cage proteins and spherical virus capsids to the fields of bionanotechnology, drug delivery, and diagnostic imaging as their internal cavities can serve as hosts for fluorophores or bioactive molecular cargo. Bacterioferritin is unusual in the ferritin protein superfamily of iron-storage cage proteins in that it contains twelve heme cofactors and is homomeric. The goal of the present study is to expand the capabilities of ferritins by developing new approaches to molecular cargo encapsulation employing bacterioferritin. Two strategies were explored to control the encapsulation of a diverse range of molecular guests compared to random entrapment, a predominant strategy employed in this area. The first was the inclusion of histidine-tag peptide fusion sequences within the internal cavity of bacterioferritin. This approach allowed for the successful and controlled encapsulation of a fluorescent dye, a protein (fluorescently labeled streptavidin), or a 5 nm gold nanoparticle. The second strategy, termed the heme-dependent cassette strategy, involved the substitution of the native heme with heme analogs attached to (i) fluorescent dyes or (ii) nickel-nitrilotriacetate (NTA) groups (which allowed for controllable encapsulation of a histidine-tagged green fluorescent protein). An in silico docking approach identified several small molecules able to replace the heme and capable of controlling the quaternary structure of the protein. A transglutaminase-based chemoenzymatic approach to surface modification of this cage protein was also accomplished, allowing for future nanoparticle targeting. This research presents novel strategies to control a diverse set of molecular encapsulations and adds a further level of sophistication to internal protein cavity engineering.

Imparting conformational memory for material adhesion.

Adhesion between similar and dissimilar materials is essential to many biological systems and synthetic materials, devices, and machines. Since the inception of adhesion science more than five decades ago, adhesion to a surface has long been recognized as beyond two-dimensional. Similarly, molecular conformation - the three-dimensional arrangement of atoms in a molecule - is ubiquitous in biology and fundamental to the binding of biomolecules. However, the connection between these concepts, which could link molecular conformation in biology to micro- and macroscopic adhesion in materials science, remains elusive. Herein, we examine this connection by manipulating the molecular conformation of a mussel-inspired universal coating, which imparts a memory for recognizing different hydrogels. This approach leads to significantly (several fold) increased interfacial adhesion between the coating and hydrogels across a broad range of length scales, from molecular to macroscopic. Furthermore, we demonstrate that imparting memory is a general and facile noncovalent approach for enhancing interfacial adhesion that, with suitable energy dissipation, can be used for the bonding of materials.

The Charge-State and Structural Stability of Peptides Conferred by Microsolvating Environments in Differential Mobility Spectrometry.

The presence of solvent vapor in a differential mobility spectrometry (DMS) cell creates a microsolvating environment that can mitigate complications associated with field-induced heating. In the case of peptides, the microsolvation of protonation sites results in a stabilization of charge density through localized solvent clustering, sheltering the ion from collisional activation. Seeding the DMS carrier gas (N2) with a solvent vapor prevented nearly all field-induced fragmentation of the protonated peptides GGG, AAA, and the Lys-rich Polybia-MP1 (IDWKKLLDAAKQIL-NH2). Modeling the microsolvation propensity of protonated n-propylamine [PrNH3]+, a mimic of the Lys side chain and N-terminus, with common gas-phase modifiers (H2O, MeOH, EtOH, iPrOH, acetone, and MeCN) confirms that all solvent molecules form stable clusters at the site of protonation. Moreover, modeling populations of microsolvated clusters indicates that species containing protonated amine moieties exist as microsolvated species with one to six solvent ligands at all effective ion temperatures (Teff) accessible during a DMS experiment (ca. 375-600 K). Calculated Teff of protonated GGG, AAA, and Polybia-MPI using a modified two-temperature theory approach were up to 86 K cooler in DMS environments seeded with solvent vapor compared to pure N2 environments. Stabilizing effects were largely driven by an increase in the ion's apparent collision cross section and by evaporative cooling processes induced by the dynamic evaporation/condensation cycles incurred in the presence of an oscillating electric separation field. When the microsolvating partner was a protic solvent, abstraction of a proton from [MP1 + 3H]3+ to yield [MP1 + 2H]2+ was observed. This result was attributed to the proclivity of protic solvents to form hydrogen-bond networks with enhanced gas-phase basicity. Collectively, microsolvation provides analytes with a solvent "air bag," whereby charge reduction and microsolvation-induced stabilization were shown to shelter peptides from the fragmentation induced by field heating and may play a role in preserving native-like ion configurations.

Design, synthesis and photocytotoxicity of upconversion nanoparticles: Potential applications for near-infrared photodynamic and photothermal therapy.

Photodynamic therapy (PDT) and photothermal therapy (PTT) are emerging modalities for the treatment of tumors and nonmalignant conditions, based on the use of photosensitizers to generate singlet oxygen or heat, respectively, upon light (laser) irradiation. They have potential advantages over conventional treatments, being minimally invasive with precise spatial-temporal selectivity and reduced side effects. However, most clinically employed PDT agents are activated at visible (vis) wavelengths for which the tissue penetration and, hence, effective treatment depth are compromised. In addition, the lipophilicity of near-infrared (NIR) photothermal agents limits their use and efficiency. To achieve combined PDT/PTT effects, both excitation wavelengths need to be tuned into the NIR spectral window of biological tissues. This paper reports the synthesis of neodymium-doped upconversion nanoparticles (NaYF4 :Yb,Er,Nd@NaYF4 :Nd) that convert 800 nm light into vis wavelengths, which can then activate conventional photosensitizers on the nanoparticle surface for PDT. Covalently bonded IR-780 dyes can readily be activated by 800 nm laser irradiation. The PEGylated nanoplatform exhibited a narrow size distribution, good stability and efficient generation of singlet oxygen under laser irradiation. The in vitro photocytotoxicity of this engineered nanoplatform as either a PDT or PTT agent in HeLa cells is demonstrated, while fluorescence microscopy in nanoplatform-incubated cells highlights its potential for bioimaging.

Squarate-based carbocyclic nucleosides: Syntheses, computational analyses and anticancer/antiviral evaluation.

Squaric acid and its derivatives are versatile synthons and have demonstrated applications in medicinal chemistry, notably as non-classical bioisosteric replacements for functional groups such as carboxylic acids, alpha-amino acids, urea, guanidine, peptide bonds and phosphate/pyrophosphate linkages. Surprisingly, no reports have appeared concerning its possible application as a nucleobase substitute in nucleosides. A preliminary investigation of such an application is reported herein. 3-Amino-4-((1R,4S)-4-(hydroxymethyl)cyclopent-2-en-1-yl)amino-cyclobut-3-ene-1,2-dione, 3-((1R,4S)-4-(hydroxymethyl)cyclopent-2-en-1-yl)amino-4-methoxycyclobut-3-ene-1,2-dione, and 3-hydroxy-4-((1R,4S)-4-(hydroxymethyl)cyclopent-2-en-1-yl)amino-cyclobut-3-ene-1,2-dione sodium salt were synthesized. Computational analyses of their structures and preliminary antitumor and antiviral screening results are reported.

Bioorthogonal Modification of the Major Sheath Protein of Bacteriophage M13: Extending the Versatility of Bionanomaterial Scaffolds.

With a mass of ∼1.6 × 107 Daltons and composed of approximately 2700 proteins, bacteriophage M13 has been employed as a molecular scaffold in bionanomaterials fabrication. In order to extend the versatility of M13 in this area, residue-specific unnatural amino acid incorporation was employed to successfully display azide functionalities on specific solvent-exposed positions of the pVIII major sheath protein of this bacteriophage. Employing a combination of engineered mutants of the gene coding for the pVIII protein, the methionine (Met) analog, l-azidohomoalanine (Aha), and a suitable Escherichia coli Met auxotroph for phage production, conditions were developed to produce M13 bacteriophage labeled with over 350 active azides (estimated by fluorescent dye labeling utilizing a strain-promoted azide-alkyne cycloaddition) and capable of azide-selective attachment to 5 nm gold nanoparticles as visualized by transmission electron microscopy. The capability of this system to undergo dual labeling utilizing both chemical acylation and bioorthogonal cycloaddition reactions was also verified. The above stratagem should prove particularly advantageous in the preparation of assemblies of larger and more complex molecular architectures based on the M13 building block.

Glyoxalase biochemistry.

The glyoxalase enzyme system utilizes intracellular thiols such as glutathione to convert α-ketoaldehydes, such as methylglyoxal, into D-hydroxyacids. This overview discusses several main aspects of the glyoxalase system and its likely function in the cell. The control of methylglyoxal levels in the cell is an important biochemical imperative and high levels have been associated with major medical symptoms that relate to this metabolite's capability to covalently modify proteins, lipids and nucleic acid.

Functional roles in S-adenosyl-L-methionine binding and catalysis for active site residues of the thiostrepton resistance methyltransferase.

Resistance to the antibiotic thiostrepton, in producing Streptomycetes, is conferred by the S-adenosyl-L-methionine (SAM)-dependent SPOUT methyltransferase Tsr. For this and related enzymes, the roles of active site amino acids have been inadequately described. Herein, we have probed SAM interactions in the Tsr active site by investigating the catalytic activity and the thermodynamics of SAM binding by site-directed Tsr mutants. Two arginine residues were demonstrated to be critical for binding, one of which appears to participate in the catalytic reaction. Additionally, evidence consistent with the involvement of an asparagine in the structural organization of the SAM binding site is presented.

Modulating glyoxalase I metal selectivity by deletional mutagenesis: underlying structural factors contributing to nickel activation profiles.

Metabolically produced methylglyoxal is a cytotoxic compound that can lead to covalent modification of cellular DNA, RNA and protein. One pathway to detoxify this compound is via the glyoxalase enzyme system. The first enzyme of this detoxification system, glyoxalase I (GlxI), can be divided into two classes according to its metal activation profile, a Zn(2+)-activated class and a Ni(2+)-activated class. In order to elucidate some of the key structural features required for selective metal activation by these two classes of GlxI, deletional mutagenesis was utilized to remove, in a step-wise fashion, a key α-helix (residues 73-87) and two small loop regions (residues 99-103 and 111-114) from the Zn(2+)-activated Pseudomonas aeruginosa GlxI (GloA3) in order to mimic the smaller Ni(2+)-activated GlxI (GloA2) from the same organism. This approach was observed to clearly shift the metal activation profile of a Zn(2+)-activated class GlxI into a Ni(2+)-activated class GlxI enzyme. The α-helix structural component was found to contribute significantly toward GlxI metal specificity, while the two small loop regions were observed to play a more crucial role in the magnitude of the enzymatic activity. The current study should provide additional information on the fundamental relationship of protein structure to metal selectivity in these metalloenzymes.

The crystal structure of a homodimeric Pseudomonas glyoxalase†I enzyme reveals asymmetric metallation commensurate with half-of-sites activity.

The Zn inactive class of glyoxalase I (Glo1) metalloenzymes are typically homodimeric with two metal-dependent active sites. While the two active sites share identical amino acid composition, this class of enzyme is optimally active with only one metal per homodimer. We have determined the X-ray crystal structure of GloA2, a Zn inactive Glo1 enzyme from Pseudomonas aeruginosa. The presented structures exhibit an unprecedented metal-binding arrangement consistent with half-of-sites activity: one active site contains a single activating Ni(2+) ion, whereas the other contains two inactivating Zn(2+) ions. Enzymological experiments prompted by the binuclear Zn(2+) site identified a novel catalytic property of GloA2. The enzyme can function as a Zn(2+) /Co(2+) -dependent hydrolase, in addition to its previously determined glyoxalase I activity. The presented findings demonstrate that GloA2 can accommodate two distinct metal-binding arrangements simultaneously, each of which catalyzes a different reaction.

Bacterial glyoxalase I enzymes: structural and biochemical investigations.

A number of bacterial glyoxalase I enzymes are maximally activated by Ni2+ and Co2+ ions, but are inactive in the presence of Zn2+, yet these enzymes will also bind this metal ion. The structure-activity relationships between these two classes of glyoxalase I serve as important clues as to how the molecular structures of these proteins control metal-activation profiles.

Molecular interactions between thiostrepton and the TipAS protein from Streptomyces lividans.

In Streptomyces lividans, the expression of several proteins is stimulated by the thiopeptide antibiotic thiostrepton. Two of these, TipAL and TipAS, autoregulate their expression after covalently binding to thiostrepton; this irreversibly sequesters the antibiotic and desensitizes the organism to its effects. In this work, additional molecular recognition interactions involved in this critical event were explored by utilizing various thiostrepton analogues and several site-directed mutants of the TipAS antibiotic binding protein. Dissociation constants for several thiostrepton analogues ranged from 0.19 to 12.95 µM, depending on the analogue. The contributions of specific structural elements of the thiostrepton molecule to this interaction have been discerned, and an unusual covalent modification between the antibiotic and a new residue in a TipAS mutant has been detected.

Ni2+-activated glyoxalase I from Escherichia coli: substrate specificity, kinetic isotope effects and evolution within the ßαßßß superfamily.

The Escherichia coli glyoxalase system consists of the metalloenzymes glyoxalase I and glyoxalase II. Little is known regarding Ni(2+)-activated E. coli glyoxalase I substrate specificity, its thiol cofactor preference, the presence or absence of any substrate kinetic isotope effects on the enzyme mechanism, or whether glyoxalase I might catalyze additional reactions similar to those exhibited by related ßαßßß structural superfamily members. The current investigation has shown that this two-enzyme system is capable of utilizing the thiol cofactors glutathionylspermidine and trypanothione, in addition to the known tripeptide glutathione, to convert substrate methylglyoxal to non-toxic D-lactate in the presence of Ni(2+) ion. E. coli glyoxalase I, reconstituted with either Ni(2+) or Cd(2+), was observed to efficiently process deuterated and non-deuterated phenylglyoxal utilizing glutathione as cofactor. Interestingly, a substrate kinetic isotope effect for the Ni(2+)-substituted enzyme was not detected; however, the proton transfer step was observed to be partially rate limiting for the Cd(2+)-substituted enzyme. This is the first non-Zn(2+)-activated GlxI where a metal ion-dependent kinetic isotope effect using deuterium-labelled substrate has been observed. Attempts to detect a glutathione conjugation reaction with the antibiotic fosfomycin, similar to the reaction catalyzed by the related superfamily member FosA, were unsuccessful when utilizing the E. coli glyoxalase I E56A mutein.

Structural variation in bacterial glyoxalase I enzymes: investigation of the metalloenzyme glyoxalase I from Clostridium acetobutylicum.

The glyoxalase system catalyzes the conversion of toxic, metabolically produced α-ketoaldehydes, such as methylglyoxal, into their corresponding nontoxic 2-hydroxycarboxylic acids, leading to detoxification of these cellular metabolites. Previous studies on the first enzyme in the glyoxalase system, glyoxalase I (GlxI), from yeast, protozoa, animals, humans, plants, and Gram-negative bacteria, have suggested two metal activation classes, Zn(2+) and non-Zn(2+) activation. Here, we report a biochemical and structural investigation of the GlxI from Clostridium acetobutylicum, which is the first GlxI enzyme from Gram-positive bacteria that has been fully characterized as to its three-dimensional structure and its detailed metal specificity. It is a Ni(2+)/Co(2+)-activated enzyme, in which the active site geometry forms an octahedral coordination with one metal atom, two water molecules, and four metal-binding ligands, although its inactive Zn(2+)-bound form possesses a trigonal bipyramidal geometry with only one water molecule liganded to the metal center. This enzyme also possesses a unique dimeric molecular structure. Unlike other small homodimeric GlxI where two active sites are located at the dimeric interface, the C. acetobutylicum dimeric GlxI enzyme also forms two active sites but each within single subunits. Interestingly, even though this enzyme possesses a different dimeric structure from previously studied GlxI, its metal activation characteristics are consistent with properties of other GlxI. These findings indicate that metal activation profiles in this class of enzyme hold true across diverse quaternary structure arrangements.

Mechanistic studies on the enzymatic processing of fluorinated methionine analogues by Trichomonas vaginalis methionine γ-lyase.

TFM (L-trifluoromethionine), a potential prodrug, was reported to be toxic towards human pathogens that express MGL (L-methionine γ-lyase; EC 4.4.1.11), a pyridoxal phosphate-containing enzyme that converts L-methionine into α-oxobutyrate, ammonia and methyl mercaptan. It has been hypothesized that the extremely reactive thiocarbonyl difluoride is produced when the enzyme acts upon TFM, resulting in cellular toxicity. The potential application of the fluorinated thiomethyl group in other areas of biochemistry and medicinal chemistry requires additional studies. Therefore a detailed investigation of the theoretical and experimental chemistry and biochemistry of these fluorinated groups (CF3S⁻ and CF2HS⁻) has been undertaken to trap and identify chemical intermediates produced by enzyme processing of molecules containing these fluorinated moieties. TvMGL (MGL from Trichomonas vaginalis) and a chemical model system of the reaction were utilized in order to investigate the cofactor-dependent activation of TFM and previously uninvestigated DFM (L-difluoromethionine). The differences in toxicity between TFM and DFM were evaluated against Escherichia coli expressing TvMGL1, as well as the intact human pathogen T. vaginalis. The relationship between the chemical structure of the reactive intermediates produced from the enzymatic processing of these analogues and their cellular toxicity are discussed.

Controllable delivery of small-molecule compounds to targeted cells utilizing carbon nanotubes.

Carbon nanotubes (CNTs) have emerged as a new alternative and efficient tool for transporting molecules with biotechnological and biomedical applications, because of their remarkable physicochemical properties. Encapsulation of functional molecules into the hollow chambers of CNTs can not only stabilize encapsulated molecules but also generate new nanodevices. In this work, we have demonstrated that CNTs can function as controllable carriers to transport small-molecule compounds (SMCs) loaded inside their hollow tunnels onto targeted cells. Using indole as model compound, CNTs can protect indole molecules during transportation. Labeling indole-loaded CNTs (indole@CNTs) with EphB4-binding peptides generates cell-homing indole@CNTs (CIDs). CIDs can selectively target EphB4-expressing cells and release indole onto cell surfaces by near-infrared (NIR) irradiation. Released indole molecules exhibit significant cell-killing effects without causing local overheating. This establishes CNTs as excellent near-infrared controllable delivery vehicles for SMCs as selective cell-killing agents.