SCPL acyltransferases catalyze the metabolism of chlorogenic acid during purple coneflower seed germination.

The metabolism of massively accumulated chlorogenic acid is crucial for the successful germination of purple coneflower (Echinacea purpurea (L.) Menoch). A serine carboxypeptidase-like (SCPL) acyltransferase (chicoric acid synthase, CAS) utilizes chlorogenic acid to produce chicoric acid during germination. However, it seems that the generation of chicoric acid lags behind the decrease in chlorogenic acid, suggesting an earlier route of chlorogenic acid metabolism. We discovered another chlorogenic acid metabolic product, 3,5-dicaffeoylquinic acid, which is produced before chicoric acid, filling the lag phase. Then, we identified two additional typical clade IA SCPL acyltransferases, named chlorogenic acid condensing enzymes (CCEs), that catalyze the biosynthesis of 3,5-dicaffeoylquinic acid from chlorogenic acid with different kinetic characteristics. Chlorogenic acid inhibits radicle elongation in a dose-dependent manner, explaining the potential biological role of SCPL acyltransferases-mediated continuous chlorogenic acid metabolism during germination. Both CCE1 and CCE2 are highly conserved among Echinacea species, supporting the observed metabolism of chlorogenic acid to 3,5-dicaffeoylquinic acid in two Echinacea species without chicoric acid accumulation. The discovery of SCPL acyltransferase involved in the biosynthesis of 3,5-dicaffeoylquinic acid suggests convergent evolution. Our research clarifies the metabolism strategy of chlorogenic acid in Echinacea species and provides more insight into plant metabolism.

Transcription elongation mechanisms of RNA polymerases I, II, and III and their therapeutic implications.

Transcription is a tightly regulated, complex, and essential cellular process in all living organisms. Transcription is comprised of three steps, transcription initiation, elongation, and termination. The distinct transcription initiation and termination mechanisms of eukaryotic RNA polymerases I, II, and III (Pols I, II, and III) have long been appreciated. Recent methodological advances have empowered high-resolution investigations of the Pols' transcription elongation mechanisms. Here, we review the kinetic similarities and differences in the individual steps of Pol I-, II-, and III-catalyzed transcription elongation, including NTP binding, bond formation, pyrophosphate release, and translocation. This review serves as an important summation of Saccharomyces cerevisiae (yeast) Pol I, II, and III kinetic investigations which reveal that transcription elongation by the Pols is governed by distinct mechanisms. Further, these studies illustrate how basic, biochemical investigations of the Pols can empower the development of chemotherapeutic compounds.

Susceptibility of Cu(I) transport ATPases to sodium dodecyl sulfate. Relevance of the composition of the micellar phase.

Sodium dodecyl sulfate (SDS) is a well-known protein denaturing agent. A less known property of this detergent is that it can activate or inactivate some enzymes at sub-denaturing concentrations. In this work we explore the effect of SDS on the ATPase activity of a hyper-thermophilic and a mesophilic Cu(I) ATPases reconstituted in mixed micelles of phospholipids and a non-denaturing detergent. An iterative procedure was used to evaluate the partition of SDS between the aqueous and the micellar phases, allowing to determine the composition of micelles prepared from phospholipid/detergent mixtures. The incubation of enzymes with SDS in the presence of different amounts of phospholipids reveals that higher SDS concentrations are required to obtain the same degree of inactivation when the initial concentration of phospholipids is increased. Remarkably, we found that, if represented as a function of the mole fraction of SDS in the micelle, the degree of inactivation obtained at different amounts of amphiphiles converges to a single inactivation curve. To interpret this result, we propose a simple model involving active and inactive enzyme molecules in equilibrium. This model allowed us to estimate the Gibbs free energy change for the inactivation process and its derivative with respect to the mole fraction of SDS in the micellar phase, the latter being a measure of the susceptibility of the enzyme to SDS. Our results showed that the inactivation free energy changes are similar for both proteins. Conversely, susceptibility to SDS is significantly lower for the hyperthermophilic ATPase, suggesting an inverse relation between thermophilicity and susceptibility to SDS.

A small-molecule PI3Kα activator for cardioprotection and neuroregeneration.

Harnessing the potential beneficial effects of kinase signalling through the generation of direct kinase activators remains an underexplored area of drug development1-5. This also applies to the PI3K signalling pathway, which has been extensively targeted by inhibitors for conditions with PI3K overactivation, such as cancer and immune dysregulation. Here we report the discovery of UCL-TRO-1938 (referred to as 1938 hereon), a small-molecule activator of the PI3Kα isoform, a crucial effector of growth factor signalling. 1938 allosterically activates PI3Kα through a distinct mechanism by enhancing multiple steps of the PI3Kα catalytic cycle and causes both local and global conformational changes in the PI3Kα structure. This compound is selective for PI3Kα over other PI3K isoforms and multiple protein and lipid kinases. It transiently activates PI3K signalling in all rodent and human cells tested, resulting in cellular responses such as proliferation and neurite outgrowth. In rodent models, acute treatment with 1938 provides cardioprotection from ischaemia-reperfusion injury and, after local administration, enhances nerve regeneration following nerve crush. This study identifies a chemical tool to directly probe the PI3Kα signalling pathway and a new approach to modulate PI3K activity, widening the therapeutic potential of targeting these enzymes through short-term activation for tissue protection and regeneration. Our findings illustrate the potential of activating kinases for therapeutic benefit, a currently largely untapped area of drug development.

Small-molecule activation of OGG1 increases oxidative DNA damage repair by gaining a new function.

Oxidative DNA damage is recognized by 8-oxoguanine (8-oxoG) DNA glycosylase 1 (OGG1), which excises 8-oxoG, leaving a substrate for apurinic endonuclease 1 (APE1) and initiating repair. Here, we describe a small molecule (TH10785) that interacts with the phenylalanine-319 and glycine-42 amino acids of OGG1, increases the enzyme activity 10-fold, and generates a previously undescribed ß,δ-lyase enzymatic function. TH10785 controls the catalytic activity mediated by a nitrogen base within its molecular structure. In cells, TH10785 increases OGG1 recruitment to and repair of oxidative DNA damage. This alters the repair process, which no longer requires APE1 but instead is dependent on polynucleotide kinase phosphatase (PNKP1) activity. The increased repair of oxidative DNA lesions with a small molecule may have therapeutic applications in various diseases and aging.

Structure-activity studies of PTPRD phosphatase inhibitors identify a 7-cyclopentymethoxy illudalic acid analog candidate for development.

Interest in development of potent, selective inhibitors of the phosphatase from the receptor type protein tyrosine phosphatase PTPRD as antiaddiction agents is supported by human genetics, mouse models and studies of our lead compound PTPRD phosphatase inhibitor, 7-butoxy illudalic acid analog 1 (7-BIA). We now report structure-activity relationships for almost 70 7-BIA-related compounds and results that nominate a 7- cyclopentyl methoxy analog as a candidate for further development. While efforts to design 7-BIA analogs with substitutions for other parts failed to yield potent inhibitors of PTPRD's phosphatase, ten 7-position substituted analogs displayed greater potency at PTPRD than 7-BIA. Several were more selective for PTPRD vs the receptor type protein tyrosine phosphatases S, F and J or the nonreceptor type protein tyrosine phosphatase N1 (PTPRS, PTPRF, PTPRJ or PTPN1/PTP1B), phosphatases at which 7-BIA displays activity. In silico studies aided design of novel analogs. A 7-position cyclopentyl methoxy substituted 7-BIA analog termed NHB1109 displayed 600-700 nM potencies in inhibiting PTPRD and PTPRS, improved selectivity vs PTPRS, PTPRF, PTPRJ or PTPN1/PTP1B phosphatases, no substantial potency at other protein tyrosine phosphatases screened, no significant potency at any of the targets of clinically-useful drugs identified in EUROFINS screens and significant oral bioavailability. Oral doses up to 200 mg/kg were well tolerated by mice, though higher doses resulted in reduced weight and apparent ileus without clear organ histopathology. NHB1109 provides a good candidate to advance to in vivo studies in addiction paradigms and toward human use to reduce reward from addictive substances.

Influence of zinc doping on the molecular biocompatibility of cadmium-based quantum dots: Insights from the interaction with trypsin.

Doping quantum dots (QDs) with extra element presents a promising future for their applications in the fields of environmental monitoring, commercial products and biomedical sciences. However, it remains unknown for the influence of doping on the molecular biocompatibility of QDs and the underlying mechanisms of the interaction between doped-QDs and protein molecules. Using the "one-pot" method, we synthesized N-acetyl-l-cysteine capped CdTe: Zn2+ QDs with higher fluorescence quantum yield, improved stability and better molecular biocompatibility compared with undoped CdTe QDs. Using digestive enzyme trypsin (TRY) as the protein model, the interactions of undoped QDs and Zn-doped QDs with TRY as well as the underlying mechanisms were investigated using multi-spectroscopy, isothermal titration calorimetry and dialysis techniques. Van der Waals forces and hydrogen bonds are the major driving forces in the interaction of both QDs with TRY, which leading to the loosening of protein skeleton and tertiary structural changes. Compared with undoped QDs, Zn-doped QDs bind less amount of TRY with a higher affinity and then release higher amount of Cd. Zn-doped QDs have a less stimulating impact on TRY activity by decreasing TRY binding and reducing Cd binding to TRY. Taken all together, Zn-doped QDs offer a safer alternative for the applications of QDs by reducing unwanted interactions with proteins and improving biocompatibility at the molecular level.

Metal ion effects on Polyphenol Oxidase Covalently immobilized on a Bio-Composite.

Biosensors can be developed using different immobilization methods. Interest in immobilization methods have increased because biosensors have been important for science. Polyphenol oxidase (PPO) was used generally in biosensor applications. For this purpose, Polyphenol oxidase from banana was purified and covalently immobilized on chitosan-gelatin bio-composite. The properties of immobilized enzyme were investigated and compared to free enzyme. Various parameters were studied such as pH, temperature and storage stability on immobilized and free enzyme. Kinetic parameters were also evaluated by different substrates on immobilized and free enzyme. Catechol was determined the best substrate for immobilized enzyme with optimum condition. In vitro effects of metal ions were studied on immobilized enzyme. Concentration range of metal ions is 1.0-10.0 x10-6 mol/L. The activity of immobilized PPO was increased by Fe+2 and Ag+1 ion. Co+3 and Cu+1 had very strong inhibitory effects with IC50 values of 19.69x10-3 mol/L and 23.49 x10-3 mol/L, respectively. Inhibition constants (Ki) and inhibition types of metal ions were determined with immobilized enzyme. Zn+2 and Cr+3 ions were showed competitive inhibition and Pb+2 ions were determined non-competitive inhibition with immobilized enzyme. Mixed type inhibition was obtained with Co+3 ion using catechol as substrate with 3.33x10-5 mol/L Ki value on immobilized PPO. Immobilized PPO can be evaluated for biosensor for the purpose of measurements of metal ions.

Inhibitory Potential of New Phenolic Hydrazide-Hydrazones with a Decoy Substrate Fragment towards Laccase from a Phytopathogenic Fungus: SAR and Molecular Docking Studies.

Laccase from pathogenic fungi participates in both the delignification and neutralization of phytoantibiotics. Furthermore, it interferes with the hormone signaling in plants and catalyzes melanization. Infections of these pathogens contribute to loss in forestry, agriculture, and horticulture. As there is still a need to expand knowledge on efficient defense strategies against phytopathogenic fungi, the present study aimed to reveal more information on the molecular mechanisms of laccase inhibition with natural and natural-like carboxylic acid semi-synthetic derivatives. A set of hydrazide-hydrazones derived from carboxylic acids, generally including electron-rich arene units that serve as a decoy substrate, was synthesized and tested with laccase from Trametes versicolor. The classic synthesis of the title inhibitors proceeded with good to almost quantitative yield. Ninety percent of the tested molecules were active in the range of KI = 8-233 µM and showed different types of action. Such magnitude of inhibition constants qualified the hydrazide-hydrazones as strong laccase inhibitors. Molecular docking studies supporting the experimental data explained the selected derivatives' interactions with the enzyme. The results are promising in developing new potential antifungal agents mitigating the damage scale in the plant cultivation, gardening, and horticulture sectors.

Enhanced whole-cell biotransformation of 3-chloropropiophenone into 1-phenyl-1-propanone by hydrogel entrapped Chlorella emersonii (211.8b).

OBJECTIVES: This study focuses on dehalogenation of halogenated organic substrate (3-Chloropropiophenone) using both free and hydrogel entrapped microalgae Chlorella emersonii (211.8b) as biocatalyst. We aimed at successful immobilization of C. emersonii (211.8b) cells and to assess their biotransformation efficiency. RESULTS: Aquasorb (entrapping material in this study) was found to be highly biocompatible with the cellular growth and viability of C. emersonii. A promising number of entrapped cells was achieved in terms of colony-forming units (CFUs = 2.1 × 104) per hydrogel bead with a comparable growth pattern to that of free cells. It was determined that there is no activity of hydrogenase that could transform 1-phenyl-2-propenone into 1-phenyl-1-propanone because after 12 h the ratio between two products (0.36 ± 0.02) remained constant throughout. Furthermore, it was found that the entrapped cells have higher biotransformation of 3-chloropropiophenone to 1-phenyl-1-propanone as compared to free cells at every interval of time. 1-phenyl-2-propenone was excluded from the whole-cell biotransformation as it was also found in the control group (due to spontaneous generation). CONCLUSION: Hence, enhanced synthesis of 1-phenyl-1-propanone by entrapped Chlorella (211.8b) can be ascribed to either an enzymatic activity (dehalogenase) or thanks to the antioxidants from 211-8b, especially when they are in immobilized form. The aquasorb based immobilization of microalgae is highly recommended as an effective tool for exploiting microalgal potentials of biocatalysis specifically when free cells activities are seized due to stress.

Covalent inhibition of P. falciparum ferredoxin-NADP+ reductase: Exploring alternative strategies for the development of new antimalarial drugs.

The protozoan Plasmodium falciparum is the main aetiological agent of tropical malaria. Characteristic of the phylum is the presence of a plastid-like organelle which hosts several homologs of plant proteins, including a ferredoxin (PfFd) and its NADPH-dependent reductase (PfFNR). The PfFNR/PfFd redox system is essential for the parasite, while mammals share no homologous proteins, making the enzyme an attractive target for novel and much needed antimalarial drugs. Based on previous findings, three chemically reactive residues important for PfFNR activity were identified: namely, the active-site Cys99, responsible for hydride transfer; Cys284, whose oxidation leads to an inactive dimeric form of the protein; and His286, which is involved in NADPH binding. These amino acid residues were probed by several residue-specific reagents and the two cysteines were shown to be promising targets for covalent inhibition. The quantitative and qualitative description of the reactivity of few compounds, including a repurposed drug, set the bases for the development of more potent and specific antimalarial leads.

From structure to ætiology: a new window on the biology of leucine-rich repeat kinase 2 and Parkinson's disease.

Since the discovery of mutations in leucine-rich repeat kinase 2 (LRRK2) as an underlying genetic cause for the development of Parkinson's disease (PD) in 2004 (Neuron 44, 601-607; Neuron 44, 595-600), and subsequent efforts to develop LRRK2 kinase inhibitors as a therapy for Parkinson's (Expert Opin. Ther. Targets 21, 751-753), elucidating the atomic resolution structure of LRRK2 has been a major goal of research into this protein. At over 250 kDa, the large size and complicated domain organisation of LRRK2 has made this a highly challenging target for structural biologists, however, a number of recent studies using both in vitro and in situ approaches (Nature 588, 344-349; Cell 182, 1508-1518.e1516; Cell 184, 3519-3527.e3510) have provided important new insights into LRRK2 structure and the complexes formed by this protein.

Laccases: Versatile Biocatalysts for the Synthesis of Heterocyclic Cores.

Laccases are multicopper oxidases that have shown a great potential in various biotechnological and green chemistry processes mainly due to their high relative non-specific oxidation of phenols, arylamines and some inorganic metals, and their high redox potentials that can span from 500 to 800 mV vs. SHE. Other advantages of laccases include the use of readily available oxygen as a second substrate, the formation of water as a side-product and no requirement for cofactors. Importantly, addition of low-molecular-weight redox mediators that act as electron shuttles, promoting the oxidation of complex bulky substrates and/or of higher redox potential than the enzymes themselves, can further expand their substrate scope, in the so-called laccase-mediated systems (LMS). Laccase bioprocesses can be designed for efficiency at both acidic and basic conditions since it is known that fungal and bacterial laccases exhibit distinct optimal pH values for the similar phenolic and aromatic amines. This review covers studies on the synthesis of five- and six-membered ring heterocyclic cores, such as benzimidazoles, benzofurans, benzothiazoles, quinazoline and quinazolinone, phenazine, phenoxazine, phenoxazinone and phenothiazine derivatives. The enzymes used and the reaction protocols are briefly outlined, and the mechanistic pathways described.

Characterization of a novel isoflavone glycoside-hydrolyzing ß-glucosidase from mangrove soil metagenomic library.

For the beneficial pharmacological properties of isoflavonoids and their related glycoconjugates, there is increasingly interest in their enzymatic conversion. In this study, a novel ß-glucosidase gene isolated from metagenomic library of mangrove sediment was cloned and overexpressed in Escherichia coli BL21(DE3). The purified recombination ß-glucosidase, designated as r-Bgl66, showed high catalytic activity for soy isoflavone glycosides. It converted soy isoflavone flour extract with the productivities of 0.87 mM/h for daidzein, 0.59 mM/h for genistein and 0.42 mM/h for glycitein. The kcat/Km values for daidzin, genistin and glycitin were 208.73, 222.37 and 288.07 mM-1 s-1, respectively. In addition, r-Bgl66 also exhibited the characteristic of glucose-tolerance, and the inhibition constant Ki was 471.4 mM. These properties make it a good candidate in the enzymatic hydrolysis of soy isoflavone glycosides. This study also highlights the utility of metagenomic approach in discovering novel ß-glucosidase for soy isoflavone glycosides hydrolysis.

PRMT1-dependent regulation of RNA metabolism and DNA damage response sustains pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer that has remained clinically challenging to manage. Here we employ an RNAi-based in vivo functional genomics platform to determine epigenetic vulnerabilities across a panel of patient-derived PDAC models. Through this, we identify protein arginine methyltransferase 1 (PRMT1) as a critical dependency required for PDAC maintenance. Genetic and pharmacological studies validate the role of PRMT1 in maintaining PDAC growth. Mechanistically, using proteomic and transcriptomic analyses, we demonstrate that global inhibition of asymmetric arginine methylation impairs RNA metabolism, which includes RNA splicing, alternative polyadenylation, and transcription termination. This triggers a robust downregulation of multiple pathways involved in the DNA damage response, thereby promoting genomic instability and inhibiting tumor growth. Taken together, our data support PRMT1 as a compelling target in PDAC and informs a mechanism-based translational strategy for future therapeutic development.Statement of significancePDAC is a highly lethal cancer with limited therapeutic options. This study identified and characterized PRMT1-dependent regulation of RNA metabolism and coordination of key cellular processes required for PDAC tumor growth, defining a mechanism-based translational hypothesis for PRMT1 inhibitors.

Acidic pH irreversibly activates the signaling enzyme SARM1.

SARM1, an executioner in axon degeneration, is an autoinhibitory NAD-consuming enzyme, composed of multiple domains. NMN and its analogs, CZ-48 and VMN, are the only known activators, which can release the inhibitory ARM domain from the enzymatic TIR domain. Here, we document that acid can also activate SARM1, even more efficiently than NMN, possibly via the protonation of the negative residues. Systematic mutagenesis revealed that a single mutation, E689Q in TIR, led to the constitutive activation of SARM1. It forms a salt bridge with R216 in the neighboring ARM, maintaining the autoinhibitory structure. Using this 'acid activation' protocol, mutation K597E was found to inhibit activation, while H685A eliminated SARM1 catalytic activity, revealing two distinct inhibitory mechanisms. The protocol has also been applied to differentiate two classes of chemical inhibitors. NAD, dHNN, disulfiram, CHAPS, and TRX-100 mainly inhibited the activation process, while nicotinamide and Tweens mainly inhibited SARM1 catalysis. Taken together, we demonstrate a new mechanism for SARM1 activation and decipher two distinct inhibitory mechanisms of SARM1.

Mapping the substrate specificity of the Plasmodium M1 and M17 aminopeptidases.

During malarial infection, Plasmodium parasites digest human hemoglobin to obtain free amino acids for protein production and maintenance of osmotic pressure. The Plasmodium M1 and M17 aminopeptidases are both postulated to have an essential role in the terminal stages of the hemoglobin digestion process and are validated drug targets for the design of new dual-target anti-malarial compounds. In this study, we profiled the substrate specificity fingerprints and kinetic behaviors of M1 and M17 aminopeptidases from Plasmodium falciparum and Plasmodium vivax, and the mouse model species, Plasmodium berghei. We found that although the Plasmodium M1 aminopeptidases share a largely similar, broad specificity at the P1 position, the P. falciparum M1 displays the greatest diversity in specificity and P. berghei M1 showing a preference for charged P1 residues. In contrast, the Plasmodium M17 aminopeptidases share a highly conserved preference for hydrophobic residues at the P1 position. The aminopeptidases also demonstrated intra-peptide sequence specificity, particularly the M1 aminopeptidases, which showed a definitive preference for peptides with fewer negatively charged intrapeptide residues. Overall, the P. vivax and P. berghei enzymes had a faster substrate turnover rate than the P. falciparum enzymes, which we postulate is due to subtle differences in structural dynamicity. Together, these results build a kinetic profile that allows us to better understand the catalytic nuances of the M1 and M17 aminopeptidases from different Plasmodium species.

PRMT5-mediated arginine methylation activates AKT kinase to govern tumorigenesis.

AKT is involved in a number of key cellular processes including cell proliferation, apoptosis and metabolism. Hyperactivation of AKT is associated with many pathological conditions, particularly cancers. Emerging evidence indicates that arginine methylation is involved in modulating AKT signaling pathway. However, whether and how arginine methylation directly regulates AKT kinase activity remain unknown. Here we report that protein arginine methyltransferase 5 (PRMT5), but not other PRMTs, promotes AKT activation by catalyzing symmetric dimethylation of AKT1 at arginine 391 (R391). Mechanistically, AKT1-R391 methylation cooperates with phosphatidylinositol 3,4,5 trisphosphate (PIP3) to relieve the pleckstrin homology (PH)-in conformation, leading to AKT1 membrane translocation and subsequent activation by phosphoinositide-dependent kinase-1 (PDK1) and the mechanistic target of rapamycin complex 2 (mTORC2). As a result, deficiency in AKT1-R391 methylation significantly suppresses AKT1 kinase activity and tumorigenesis. Lastly, we show that PRMT5 inhibitor synergizes with AKT inhibitor or chemotherapeutic drugs to enhance cell death. Altogether, our study suggests that R391 methylation is an important step for AKT activation and its oncogenic function.

Design, synthesis, molecular docking, and in vitro α-glucosidase inhibitory activities of novel 3-amino-2,4-diarylbenzo[4,5]imidazo[1,2-a]pyrimidines against yeast and rat α-glucosidase.

In an attempt to find novel, potent α-glucosidase inhibitors, a library of poly-substituted 3-amino-2,4-diarylbenzo[4,5]imidazo[1,2-a]pyrimidines 3a-ag have been synthesized through heating a mixture of 2-aminobenzimidazoles 1 and α-azidochalcone 2 under the mild conditions. This efficient, facile protocol has been resulted into the desirable compounds with a wide substrate scope in good to excellent yields. Afterwards, their inhibitory activities against yeast α-glucosidase enzyme were investigated. Showing IC50 values ranging from 16.4 ± 0.36 µM to 297.0 ± 1.2 µM confirmed their excellent potency to inhibit α-glucosidase which encouraged us to perform further studies on α-glucosidase enzymes obtained from rat as a mammal source. Among various synthesized 3-amino-2,4-diarylbenzo[4,5]imidazo[1,2-a]pyrimidines, compound 3k exhibited the highest potency against both Saccharomyces cerevisiae α-glucosidase (IC50 = 16.4 ± 0.36 µM) and rat small intestine α-glucosidase (IC50 = 45.0 ± 8.2 µM). Moreover, the role of amine moiety on the observed activity was studied through substituting with chlorine and hydrogen resulted into a considerable deterioration on the inhibitory activity. Kinetic study and molecular docking study have confirmed the in-vitro results.

Conformational plasticity of the ULK3 kinase domain.

The human protein kinase ULK3 regulates the timing of membrane abscission, thus being involved in exosome budding and cytokinesis. Herein, we present the first high-resolution structures of the ULK3 kinase domain. Its unique features are explored against the background of other ULK kinases. An inhibitor fingerprint indicates that ULK3 is highly druggable and capable of adopting a wide range of conformations. In accordance with this, we describe a conformational switch between the active and an inactive ULK3 conformation, controlled by the properties of the attached small-molecule binder. Finally, we discuss a potential substrate-recognition mechanism of the full-length ULK3 protein.