Exogenous Small Molecules Research Articles

The glycolytic metabolite methylglyoxal covalently inactivates the NLRP3 inflammasome

The NLRP3 inflammasome promotes inflammation in disease, yet the full repertoire of mechanisms regulating its activity is not well delineated. Among established regulatory mechanisms, covalent modification of NLRP3 has emerged as a common route for the pharmacological inactivation of this protein. Here, we show that inhibition of the glycolytic enzyme phosphoglycerate kinase 1 (PGK1) results in the accumulation of methylglyoxal, a reactive metabolite whose increased levels decrease NLRP3 assembly and inflammatory signaling in cells. We find that methylglyoxal inactivates NLRP3 via a non-enzymatic, covalent-crosslinking-based mechanism, promoting inter- and intraprotein MICA (methyl imidazole crosslink between cysteine and arginine) posttranslational linkages within NLRP3. This work establishes NLRP3 as capable of sensing a host of electrophilic chemicals, both exogenous small molecules and endogenous reactive metabolites, and suggests a mechanism by which glycolytic flux can moderate the activation status of a central inflammatory signaling pathway.

Paradoxical action of PP2A inhibition and its potential for therapeutic sensitization.

The protein phosphatase 2A (PP2A), a serine/threonine phosphatase, is recognized as a tumor suppressor involved in diverse cellular processes and essential for maintaining cell viability in vivo. However, endogenous inhibitors of PP2A such as cancerous inhibitor of PP2A (CIP2A) and endogenous nuclear protein inhibitor 2 of PP2A (SET) counteract the anticancer function of PP2A, promoting tumorigenesis, development, and drug resistance in tumors. Surprisingly though, contrary to conventional understanding, inhibition of the tumor suppressor gene PP2A with exogenous small molecule compounds can enhance the efficacy of cancer treatment and achieve superior tumor inhibition. Moreover, exogenous PP2A inhibitors resensitize cancers to treatment and provide novel therapeutic strategies for drug-resistant tumors, which warrant further investigation.

Gap-Δenergy, a New Metric of the Bond Energy State, Assisting to Predict Molecular Toxicity.

Molecular toxicity is a critical feature of drug development. It is thus very important to develop computational models to evaluate the toxicity of small molecules. The accuracy of toxicity prediction largely depends on the quality of molecular representation; however, current methods for this purpose do not address this issue well. Here, we introduce a new metric, gap-Δenergy, which is designed to quantify the intermolecular bond energy difference with atom distance. We next find significant variations in the gap-Δenergy distribution among different types of molecules. Moreover, we show that this metric is able to distinguish the toxic small molecules. We collected data sets of toxic and exogenous small molecules and presented a novel index, namely, global toxicity, to evaluate the overall toxicity of molecules. Based on molecular descriptors and the proposed gap-Δenergy metric, we further constructed machine learning models that were trained with 7816 small molecules. The XGBoost-based model achieved the best performance with an AUC score of 0.965 and an F1 score of 0.849 on the test set (1954 small molecules), which outperformed the model that did not use gap-Δenergy features, with a sensitivity score increase of 3.2%.

Orthogonal inducible control of Cas13 circuits enables programmable RNA regulation in mammalian cells.

RNA plays an indispensable role in mammalian cell functions. Cas13, a class of RNA-guided ribonuclease, is a flexible tool for modifying and regulating coding and non-coding RNAs, with enormous potential for creating new cell functions. However, the lack of control over Cas13 activity has limited its cell engineering capability. Here, we present the CRISTAL (Control of RNA with Inducible SpliT CAs13 Orthologs and Exogenous Ligands) platform. CRISTAL is powered by a collection (10 total) of orthogonal split inducible Cas13 effectors that can be turned ON or OFF via small molecules in multiple cell types, providing precise temporal control. Also, we engineer Cas13 logic circuits that can respond to endogenous signaling and exogenous small molecule inputs. Furthermore, the orthogonality, low leakiness, and high dynamic range of our inducible Cas13d and Cas13b enable the design and construction of a robust incoherent feedforward loop, leading to near-perfect and tunable adaptation response. Finally, using our inducible Cas13 effectors, we achieve simultaneous multiplexed control of multiple genes in vitro and in mice. Together, our CRISTAL design represents a powerful platform for precisely regulating RNA dynamics to advance cell engineering and elucidate RNA biology.

Identification of novel F2-isoprostane metabolites by specific UDP-glucuronosyltransferases

UDP-glucuronosyltransferases (UGTs) catalyze the conjugation of glucuronic acid with endogenous and exogenous lipophilic small molecules to facilitate their inactivation and excretion from the body. This represents approximately 35 % of all phase II metabolic transformations. Fatty acids and their oxidized eicosanoid derivatives can be metabolized by UGTs. F2-isoprostanes (F2-IsoPs) are eicosanoids formed from the free radical oxidation of arachidonic acid. These molecules are potent vasoconstrictors and are widely used as biomarkers of endogenous oxidative damage. An increasing body of evidence demonstrates the efficacy of measuring the β-oxidation metabolites of F2-IsoPs rather than the unmetabolized F2-IsoPs to quantify oxidative damage in certain settings. Yet, the metabolism of F2-IsoPs is incompletely understood. This study sought to identify and characterize novel phase II metabolites of 15-F2t-IsoP and 5-epi-5-F2t-IsoP, two abundantly produced F2-IsoPs, in human liver microsomes (HLM). Utilizing liquid chromatography-mass spectrometry, we demonstrated that glucuronide conjugates are the major metabolites of these F2-IsoPs in HLM. Further, we showed that these molecules are metabolized by specific UGT isoforms. 15-F2t-IsoP is metabolized by UGT1A3, 1A9, and 2B7, while 5-epi-5-F2t-IsoP is metabolized by UGT1A7, 1A9, and 2B7. We identified, for the first time, the formation of intact glucuronide F2-IsoPs in human urine and showed that F2-IsoP glucuronidation is reduced in people supplemented with eicosapentaenoic and docosahexaenoic acids for 12 weeks. These studies demonstrate that endogenous F2-IsoP levels can be modified by factors other than redox mechanisms.

The Role of Macrophage Migration Inhibitory Factor (MIF) and D-Dopachrome Tautomerase (D-DT/MIF-2) in Infections: A Clinical Perspective.

Macrophage migration inhibitory factor (MIF) and its homolog, D-dopachrome tautomerase (D-DT), are cytokines that play critical roles in the immune response to various infectious diseases. This review provides an overview of the complex involvement of MIF and D-DT in bacterial, viral, fungal, and parasitic infections. The role of MIF in different types of infections is controversial, as it has either a protective function or a host damage-enhancing function depending on the pathogen. Depending on the specific role of MIF, different therapeutic options for MIF-targeting drugs arise. Human MIF-neutralizing antibodies, anti-parasite MIF antibodies, small molecule MIF inhibitors or MIF-blocking peptides, as well as the administration of exogenous MIF or MIF activity-augmenting small molecules have potential therapeutic applications and need to be further explored in the future. In addition, MIF has been shown to be a potential biomarker and therapeutic target in sepsis. Further research is needed to unravel the complexity of MIF and D-DT in infectious diseases and to develop personalized therapeutic approaches targeting these cytokines. Overall, a comprehensive understanding of the role of MIF and D-DT in infections could lead to new strategies for the diagnosis, treatment, and management of infectious diseases.

B-cell lymphoma-2 family proteins in the crosshairs: Small molecule inhibitors and activators for cancer therapy.

The B-cell lymphoma-2 (BCL-2) family of proteins plays a crucial role in the regulation of apoptosis, offering a dual mechanism for its control. Numerous studies have established a strong association between gene disorders of these proteins and the proliferation of diverse cancer cell types. Consequently, the identification and development of drugs targeting BCL-2 family proteins have emerged as a prominent area in antitumor therapy. Over the last two decades, several small-molecules have been designed to modulate the protein-protein interactions between anti- and proapoptotic BCL-2 proteins, effectively suppressing tumor growth and metastasis in vivo. The primary focus of research has been on developing BCL-2 homology 3 (BH3) mimetics to target antiapoptotic BCL-2 proteins, thereby competitively releasing proapoptotic BCL-2 proteins and restoring the blocked intrinsic apoptotic program. Additionally, for proapoptotic BCL-2 proteins, exogenous small molecules have been explored to activate cell apoptosis by directly interacting with executioner proteins such as BCL-2-associated X protein (BAX) or BCL-2 homologous antagonist/killer protein (BAK). In this comprehensive review, we summarize the inhibitors and activators (sensitizers) of BCL-2 family proteins developed over the past decades, highlighting their discovery, optimization, preclinical and clinical status, and providing an overall landscape of drug development targeting these proteins for therapeutic purposes.

Wearable Microneedle-Based Array Patches for Continuous Electrochemical Monitoring and Drug Delivery: Toward a Closed-Loop System for Methotrexate Treatment.

Wearable devices based on microneedle (MN) technology have recently emerged as tools for in situ transdermal sensing or delivery in interstitial fluid (ISF). Particularly, MN-based electrochemical sensors allow the continuous monitoring of analytes in a minimally invasive manner through ISF. Exogenous small molecules found in ISF such as therapeutic drugs are ideal candidates for MN sensors due to their correlation with blood levels and their relevance for the optimal management of personalized therapies. Herein, a hollow MN array patch is modified with conductive pastes and functionalized with cross-linked chitosan to develop an MN-based voltammetric sensor for continuous monitoring of methotrexate (MTX). Interestingly, the chitosan coating avoids biofouling while enabling the adsorption of MTX at the electrode's surface for sensitive analysis. The MN sensor exhibits excellent analytical performance in vitro with protein-enriched artificial ISF and ex vivo under a Franz diffusion cell configuration. The MN sensor shows a linear range from 25 to 400 μM, which fits within the therapeutic range of high-dose MTX treatment for cancer patients and an excellent continuous operation for more than two days. Moreover, an iontophoretic hollow MN array patch is developed with the integration of both the anode and cathode in the single MN array patch. The ex vivo characterization demonstrates the transdermal on-demand drug delivery of MTX. Overall, the combination of both MN patches represents impactful progress in closed-loop systems for therapeutic drug management in disorders such as cancer, rheumatoid arthritis, or psoriasis.

In-Cell Engineering of Protein Crystals into Hybrid Solid Catalysts for Artificial Photosynthesis.

The emergence of protein-based crystalline materials offers promising opportunities in enzyme immobilization. However, the current systems used for encapsulation of protein crystals are limited to either exogenous small molecules or monomeric proteins. In this work, polyhedra crystals were used to simultaneously encapsulate the foreign enzymes FDH and the organic photocatalyst eosin Y. These hybrid protein crystals are prepared easily by cocrystallization within a cell without a requirement for complex purification processes because they spontaneously form 1 μm scale solid particles. After immobilization within protein crystals, the recombinant FDH is recyclable and thermally stable and maintains 94.4% activity compared to the free enzyme. In addition, the incorporation of eosin Y endows the solid catalyst with CO2-formate conversion activity based on a cascade reaction. This work indicates that engineering protein crystals by both in vivo and in vitro strategies will provide robust and environmentally friendly solid catalysts for artificial photosynthesis.

Recent advances in understanding brain cancer metabolomics: a review.

Regardless of the significant progress made in surgical techniques and adjuvant therapies, brain tumors are a major contributor to cancer-related morbidity and mortality in both pediatric and adult populations. Gliomas represent a significant proportion of cerebral neoplasms, exhibiting diverse levels of malignancy. The etiology and mechanisms of resistance of this malignancy are inadequately comprehended, and the optimization of patient diagnosis and prognosis is a challenge due to the diversity of the disease and the restricted availability of therapeutic options. Metabolomics refers to the comprehensive analysis of endogenous and exogenous small molecules, both in a targeted and untargeted manner, that enables the characterization of an individual's phenotype and offers valuable insights into cellular activity, particularly in the context of cancer biology, including brain tumor biology. Metabolomics has garnered attention in current years due to its potential to facilitate comprehension of the dynamic spatiotemporal regulatory network of enzymes and metabolites that enables cancer cells to adapt to their environment and foster the development of tumors. Metabolic changes are widely acknowledged as a significant characteristic for tracking the advancement of diseases, treatment efficacy, and identifying novel molecular targets for successful medical management. Metabolomics has emerged as an exciting area for personalized medicine and drug discovery, utilizing advanced analytical techniques such as nuclear magnetic resonance spectroscopy (MRS) and mass spectrometry (MS) to achieve high-throughput analysis. This review examines and highlights the latest developments in MRS, MS, and other technologies in studying human brain tumor metabolomics.

Direct tests of cytochrome c and c1 functions in the electron transport chain of malaria parasites

The mitochondrial electron transport chain (ETC) of Plasmodium malaria parasites is a major antimalarial drug target, but critical cytochrome(cyt) functions remain unstudied and enigmatic. Parasites express two distinct cyt c homologs (c and c-2) with unusually sparse sequence identity and uncertain fitness contributions. P. falciparum cyt c-2 is the most divergent eukaryotic cyt c homolog currently known and has sequence features predicted to be incompatible with canonical ETC function. We tagged both cyt c homologs and the related cyt c1 for inducible knockdown. Translational repression of cyt c and cyt c1 was lethal to parasites, which died from ETC dysfunction and impaired ubiquinone recycling. In contrast, cyt c-2 knockdown or knockout had little impact on blood-stage growth, indicating that parasites rely fully on the more conserved cyt c for ETC function. Biochemical and structural studies revealed that both cyt c and c-2 are hemylated by holocytochrome c synthase, but UV-vis absorbance and EPR spectra strongly suggest that cyt c-2 has an unusually open active site in which heme is stably coordinated by only a single axial amino acid ligand and can bind exogenous small molecules. These studies provide a direct dissection of cytochrome functions in the ETC of malaria parasites and identify a highly divergent Plasmodium cytochrome c with molecular adaptations that defy a conserved role in eukaryotic evolution.

In vivo solid phase microextraction for therapeutic monitoring and pharmacometabolomic fingerprinting of lung during in vivo lung perfusion of FOLFOX

In vivo lung perfusion (IVLP) is a novel isolated lung technique developed to enable the local, in situ administration of high-dose chemotherapy to treat metastatic lung cancer. Combination therapy using folinic acid (FOL), 5-fluorouracil (F), and oxaliplatin (OX) (FOLFOX) is routinely employed to treat several types of solid tumours in various tissues. However, F is characterized by large interpatient variability with respect to plasma concentration, which necessitates close monitoring during treatments using of this compound. Since plasma drug concentrations often do not reflect tissue drug concentrations, it is essential to utilize sample-preparation methods specifically suited to monitoring drug levels in target organs. In this work, in vivo solid-phase microextraction (in vivo SPME) is proposed as an effective tool for quantitative therapeutic drug monitoring of FOLFOX in porcine lungs during pre-clinical IVLP and intravenous (IV) trials. The concomitant extraction of other endogenous and exogenous small molecules from the lung and their detection via liquid chromatography coupled to high resolution mass spectrometry (LC-HRMS) enabled an assessment of FOLFOX's impact on the metabolomic profile of the lung and revealed the metabolic pathways associated with the route of administration (IVLP vs. IV) and the therapy itself. This study also shows that the immediate instrumental analysis of metabolomic samples is ideal, as long-term storage at −80 °C results in changes in the metabolite content in the sample extracts.

Structural Elucidation of a Polypeptoid Chain in a Crystalline Lattice Reveals Key Morphology-Directing Role of the N-Terminus.

The ability to engineer synthetic polymers with the same structural precision as biomacromolecules is crucial to enable the de novo design of robust nanomaterials with biomimetic function. Peptoids, poly(N-substituted) glycines, are a highly controllable bio-inspired polymer family that can assemble into a variety of functional, crystalline nanostructures over a wide range of sequences. Extensive investigation on the molecular packing in these lattices has been reported; however, many key atomic-level details of the molecular structure remain underexplored. Here, we use cryo-TEM 3D reconstruction to directly visualize the conformation of an individual polymer chain within a peptoid nanofiber lattice in real space at 3.6 Å resolution. The backbone in the N-decylglycine hydrophobic core is shown to clearly adopt an extended, all-cis-sigma strand conformation, as previously suggested in many peptoid lattice models. We also show that packing interactions (covalent and noncovalent) at the solvent-exposed N-termini have a dominant impact on the local chain ordering and hence the ability of the chains to pack into well-ordered lattices. Peptoids in pure water form fibers with limited growth in the a direction (<14 molecules in width), whereas in the presence of formamide, they grow to over microns in length in the a direction. This dependence points to the significant role of the chain terminus in determining the long-range order in the packing of peptoid lattices and provides an opportunity to modulate lattice stability and nanoscale morphology by the addition of exogenous small molecules. These findings help resolve a major challenge in the de novo structure-based design of sequence-defined biomimetic nanostructures based on crystalline domains and should accelerate the design of functional nanostructures.

Prediction of Fluorophore Brightness in Designed Mini Fluorescence Activating Proteins

The de novo computational design of proteins with predefined three-dimensional structure is becoming much more routine due to advancements both in force fields and algorithms. However, creating designs with functions beyond folding is more challenging. In that regard, the recent design of small beta barrel proteins that activate the fluorescence of an exogenous small molecule chromophore (DFHBI) is noteworthy. These proteins, termed mini fluorescence activating proteins (mFAPs), have been shown to increase the brightness of the chromophore more than 100-fold upon binding to the designed ligand pocket. The design process created a large library of variants with different brightness levels but gave no rational explanation for why one variant was brighter than another. Here, we use quantum mechanics and molecular dynamics simulations to investigate how molecular flexibility in the ground and excited states influences brightness. We show that the ability of the protein to resist dihedral angle rotation of the chromophore is critical for predicting brightness. Our simulations suggest that the mFAP/DFHBI complex has a rough energy landscape, requiring extensive ground-state sampling to achieve converged predictions of excited-state kinetics. While computationally demanding, this roughness suggests that mFAP protein function can be enhanced by reshaping the energy landscape toward conformations that better resist DFHBI bond rotation.

The lightweight spherical samplers for simplified collection, storage, and ambient ionization of drugs from saliva and blood

The continuous monitoring of health conditions before, during, and after spaceflight is an important task in gravitational physiology. Thus a simple, non-invasive method for biological samples collection is required, but the specific limitations, such as weight and volume restrictions, limit the variety of suitable samplers. The physiological changes exist at microgravity (μG) conditions and could influence drug pharmacokinetics. Here we present the application of lightweight spherical sampler probes for the simplified sampling of human saliva with further detection of exogenous small molecules by the ambient mass spectrometry. The rigidity and spherical form of the sampler probes allow collecting saliva smears or biological fluids without additional instruments and improve the stability of the ionization process leading to the increased sensitivity of the analysis. The detection and quantitation of acetaminophen, which is used as a model substance, was proved to demonstrate the analytical capabilities of the proposed method to the saliva smears or whole blood samples even after long-term storage at room temperature. The further investigation of drug pharmacokinetic at μG conditions might be provided by means of proposed samplers.

Interactions between Human Serum Albumin and Sulfadimethoxine Determined Using Spectroscopy and Molecular Docking.

Sulfonamides are widely used antibiotics in agricultural production. However, the potential threat of these drugs to human health has increased global concern. Human serum albumin (HSA) is the main reservoir and transporter of exogenous small molecules in humans. In this study, the interaction between sulfadimethoxine (SMT) and human serum albumin (HSA) was studied using spectroscopy and computer simulation. Our results showed that the hydrogen bonding and van der Waals forces drove SMT to enter the binding site I of HSA spontaneously and resulted in the fluorescence quenching of HSA. The stability of the HSA–SMT complex decreased with an increase in temperature. The binding of SMT to HSA induced alterations in the secondary structure of HSA, where the content of α-helix decreased from 61.0% of the free state to 59.0% of the compound state. The π–π, π–σ, and π–alkyl interactions between HSA and SMT were found to play important roles in maintaining the stability of the complex.

The Expression Profiles and Deregulation of UDP-Glycosyltransferase (UGT) Genes in Human Cancers and Their Association with Clinical Outcomes.

Simple SummaryThe human UDP-glycosyltransferase (UGT) superfamily plays a critical role in the metabolism of numerous endogenous and exogenous small lipophilic compounds, including carcinogens, drugs, and bioactive molecules with pro- or anti-cancer activity. Previous studies have documented the expression of UGT genes in several cancers derived from drug-metabolizing organs (e.g., liver, colon, kidney). The present study represents the first to comprehensively assess the expression profiles of UGT genes and their impact on patient survival in nearly 30 different cancers primarily derived from non-drug-metabolizing organs. Briefly, our comprehensive analysis of the transcriptomic (RNAseq) and clinical datasets of 9514 patients from 33 different cancers shows the widespread expression of UGT genes, indicative of active drug metabolism within the tumor through the UGT conjugation pathway. We further identified the UGT genes whose intratumoral expression was associated with patient survival, highlighting the potential of UGT genes as prognostic biomarkers and therapeutic targets in various cancers. The human UDP-glycosyltransferase (UGTs) superfamily has 22 functional enzymes that play a critical role in the metabolism of small lipophilic compounds, including carcinogens, drugs, steroids, lipids, fatty acids, and bile acids. The expression profiles of UGT genes in human cancers and their impact on cancer patient survival remains to be systematically investigated. In the present study, a comprehensive analysis of the RNAseq and clinical datasets of 9514 patients from 33 different TCGA (the Genome Cancer Atlas) cancers demonstrated cancer-specific UGT expression profiles with high interindividual variability among and within individual cancers. Notably, cancers derived from drug metabolizing tissues (liver, kidney, gut, pancreas) expressed the largest number of UGT genes (COAD, KIRC, KIRP, LIHC, PAAD); six UGT genes (1A6, 1A9, 1A10, 2A3, 2B7, UGT8) showed high expression in five or more different cancers. Kaplan–Meier plots and logrank tests revealed that six UGT genes were significantly associated with increased overall survival (OS) rates [UGT1A1 (LUSC), UGT1A6 (ACC), UGT1A7 (ACC), UGT2A3 (KIRC), UGT2B15 (BLCA, SKCM)] or decreased OS rates [UGT2B15 (LGG), UGT8 (UVM)] in specific cancers. Finally, differential expression analysis of 611 patients from 12 TCGA cancers identified 16 UGT genes (1A1, 1A3, 1A6, 1A7, 1A8, 1A9, 1A10, 2A1, 2A3, 2B4, 2B7, 2B11, 2B15, 3A1, 3A2, UGT8) that were up/downregulated in at least one cancer relative to normal tissues. In conclusion, our data show widespread expression of UGT genes in cancers, highlighting the capacity for intratumoural drug metabolism through the UGT conjugation pathway. The data also suggests the potentials for specific UGT genes to serve as prognostic biomarkers or therapeutic targets in cancers.

The Early Adhesion Effects of Human Gingival Fibroblasts on Bovine Serum Albumin Loaded Hydrogenated Titanium Nanotube Surface

The soft tissue sealing at the transmucal portion of implants is vital for the long-term stability of implants. Hydrogenated titanium nanotubes (H2-TNTs) as implant surface treatments were proved to promote the adhesion of human gingival fibroblasts (HGFs) and have broad usage as drug delivery systems. Bovine serum albumin (BSA) as the most abundant albumin in body fluid was crucial for cell adhesion and was demonstrated as a normal loading protein. As the first protein arriving on the surface of the implant, albumin plays an important role in initial adhesion of soft tissue cells, it is also a common carrier, transferring and loading different endogenous and exogenous substances, ions, drugs, and other small molecules. The aim of the present work was to investigate whether BSA-loaded H2-TNTs could promote the early adhesion of HGFs; H2-TNTs were obtained by hydrogenated anodized titanium dioxide nanotubes (TNTs) in thermal treatment, and BSA was loaded in the nanotubes by vacuum drying; our results showed that the superhydrophilicity of H2-TNTs is conducive to the loading of BSA. In both hydrogenated titanium nanotubes and non-hydrogenated titanium nanotubes, a high rate of release was observed over the first hour, followed by a period of slow and sustained release; however, BSA-loading inhibits the early adhesion of human gingival fibroblasts, and H2-TNTs has the best promoting effect on cell adhesion. With the release of BSA after 4 h, the inhibitory effect of BSA on cell adhesion was weakened.

Into the Labyrinth of the Lipocalin α1-Acid Glycoprotein.

α1-acid glycoprotein (AGP), also known as Orosomucoid (ORM), belongs to the Lipocalin protein family and it is well-known for being a positive acute-phase protein. AGP is mostly found in plasma, with the liver as main contributor, but it is also expressed in other tissues such as the brain or the adipose tissue. Despite the vast literature on AGP, the physiological functions of the protein remain to be elucidated. A large number of activities mostly related to protection and immune system modulation have been described. Recently created AGP-knockout models have suggested novel physiological roles of AGP, including regulation of metabolism. AGP has an outstanding ability to efficiently bind endogenous and exogenous small molecules that together with the complex and variable glycosylation patterns, determine AGP functions. This review summarizes and discusses the recent findings on AGP structure (including glycans), ligand-binding ability, regulation, and physiological functions of AGP. Moreover, this review explores possible molecular and functional connections between AGP and other members of the Lipocalin protein family.

Human METTL7B is an alkyl thiol methyltransferase that metabolizes hydrogen sulfide and captopril

Methylation of alkyl thiols is a biotransformation pathway designed to reduce thiol reactivity and potential toxicity, yet the gene and protein responsible for human alkyl thiol methyltransferase (TMT) activity remain unknown. Here we demonstrate with a range of experimental approaches using cell lines, in vitro systems, and recombinantly expressed enzyme, that human methyltransferase-like protein 7B (METTL7B) catalyzes the transfer of a methyl group from S-adenosyl-l-methionine (AdoMet) to hydrogen sulfide (H2S) and other exogenous thiol small molecules. METTL7B gene modulation experiments, including knockdown in HepG2 cells and overexpression in HeLa cells, directly alter the methylation of the drug captopril, a historic probe substrate for TMT activity. Furthermore, recombinantly expressed and purified wild-type METTL7B methylates several thiol compounds, including H2S, 7α-thiospironolactone, l-penicillamine, and captopril, in a time- and concentration-dependent manner. Typical for AdoMet-dependent small molecule methyltransferases, S-adenosyl-l-homocysteine (AdoHcy) inhibited METTL7B activity in a competitive fashion. Similarly, mutating a conserved aspartate residue, proposed to anchor AdoMet into the active site, to an alanine (D98A) abolished methylation activity. Endogenous thiols such as glutathione and cysteine, or classic substrates for other known small molecule S-, N-, and O-methyltransferases, were not substrates for METTL7B. Our results confirm, for the first time, that METTL7B, a gene implicated in multiple disease states including rheumatoid arthritis and breast cancer, encodes a protein that methylates small molecule alkyl thiols. Identifying the catalytic function of METTL7B will enable future pharmacological research in disease pathophysiology where altered METTL7B expression and, potentially H2S levels, can disrupt cell growth and redox state.