Enzymatic Fluoromethylation Enabled by the S-Adenosylmethionine Analog Te-Adenosyl-L-(fluoromethyl)homotellurocysteine
Ontology highlight
ABSTRACT:
SUBMITTER: Neti S
PROVIDER: S-EPMC10214534 | biostudies-literature | 2023 May
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:S-adenosylmethionine-dependent methyltransferases are involved in countless biological processes, including signal transduction, epigenetics, natural product biosynthesis, and detoxification. Only a handful of carboxylate methyltransferases have evolved to participate in amide bond formation. In this report we show that enzyme-catalyzed F-methylation of carboxylate substrates produces F-methyl esters that readily react with N- or S-nucleophiles under physiological conditions. We demonstrate the applicability of this approach to the synthesis of small amides, hydroxamates, and thioesters, as well as to site-specific protein modification and native chemical ligation.
| S-EPMC10952888 | biostudies-literature
Project description: Not available
| S-EPMC10952241 | biostudies-literature
Project description:Protein methyltransferases (PMTs) utilize S-adenosyl-L-methionine (SAM) as a cofactor and transfer its sulfonium methyl moiety to diverse substrates. These methylation events can lead to meaningful biological outcomes, from transcriptional activation/silencing to cell cycle regulation. This article describes recently developed technology based on protein engineering in tandem with SAM analog cofactors and bioorthogonal click chemistry to unambiguously profile the substrates of a specific PMT. The protocols encapsulate the logic and methods of selectively profiling the substrates of a candidate PMT by (1) engineering the selected PMT to accommodate a bulky SAM analog; (2) generating a proteome containing the engineered PMT; (3) visualizing the proteome-wide substrates of the designated PMT via bioorthogonal labeling with a fluorescent tag; and finally (4) pulling down the proteome-wide substrates for mass spectrometric analysis.
| S-EPMC3647616 | biostudies-literature
Project description:Radical S-adenosyl-l-methionine (SAM) enzymes initiate biological radical reactions with the 5'-deoxyadenosyl radical (5'-dAdo•). A [4Fe-4S]+ cluster reductively cleaves SAM to form the Ω organometallic intermediate in which the 5'-deoxyadenosyl moiety is directly bound to the unique iron of the [4Fe-4S] cluster, with subsequent liberation of 5'-dAdo•. Here we present synthesis of the SAM analog S-adenosyl-l-ethionine (SAE) and show SAE is a mechanistically-equivalent SAM-alternative for HydG, both supporting enzymatic turnover of substrate tyrosine and forming the organometallic intermediate Ω. Photolysis of SAE bound to HydG forms an ethyl radical trapped in the active site. The ethyl radical withstands prolonged storage at 77 K and its EPR signal is only partially lost upon annealing at 100 K, making it significantly less reactive than the methyl radical formed by SAM photolysis. Upon annealing above 77K, the ethyl radical adds to the [4Fe-4S]2+ cluster, generating an ethyl-[4Fe-4S]3+ organometallic species termed ΩE.
| S-EPMC8081114 | biostudies-literature
Project description:The Zika virus (ZIKV) has emerged as a major health hazard. We present here a high resolution structure (1.55 Å) of ZIKV NS5 methyltransferase bound to a novel S-adenosylmethionine (SAM) analog in which a 4-fluorophenyl moiety substitutes for the methyl group. We show that the 4-fluorophenyl moiety extends into a portion of the RNA binding tunnel that typically contains the adenosine 2'OH of the RNA-cap moiety. Together, the new SAM analog and the high-resolution crystal structure are a step towards the development of antivirals against ZIKV and other flaviviruses.
| S-EPMC5431627 | biostudies-literature
Project description:Continuous glucose monitoring from sweat and tears can improve the quality of life of diabetic patients and provide data for more accurate diagnosis and treatment. Current continuous glucose sensors use enzymes with a one-to-two week lifespan, which forces periodic replacement. Metal oxide sensors are an alternative to enzymatic sensors with a longer lifetime. However, metal oxide sensors do not operate in sweat and tears because they function at high pH (pH > 10), and sweat and tears are neutral (pH = 7). Here, we introduce a non-enzymatic metal oxide glucose sensor that functions in neutral fluids by electronically inducing a reversible and localized pH change. We demonstrate glucose monitoring at physiologically relevant levels in neutral fluids mimicking sweat, and wireless communication with a personal computer via an integrated circuit board.
| S-EPMC6659689 | biostudies-literature
Project description:The hydrolysis of nucleotides is of paramount importance as an energy source for cellular processes. In addition, the transfer of phosphates from nucleotides onto proteins is important as a post-translational protein modification. Monitoring the enzymatic turnover of nucleotides therefore offers great potential as a tool to follow enzymatic activity. While a number of fluorescence sensors are known, so far, there are no methods available for the real-time monitoring of ATP hydrolysis inside live cells. We present the synthesis and application of a novel fluorogenic adenosine 5'-tetraphosphate (Ap4) analog suited for this task. Upon enzymatic hydrolysis, the molecule displays an increase in fluorescence intensity, which provides a readout of its turnover. We demonstrate how this can be used for monitoring cellular processes involving Ap4 hydrolysis. To this end, we visualized the enzymatic activity in live cells using confocal fluorescence microscopy of the Ap4 analog. Our results demonstrate that the Ap4 analog is hydrolyzed in lysosomes. We show that this approach is suited to visualize the lysosome distribution profiles within the live cell and discuss how it can be employed to gather information regarding autophagic flux.
| S-EPMC8395338 | biostudies-literature
Project description:S-Adenosyl methionine (SAM)-dependent methyltransferases hold significant potential as tools for the biocatalytic synthesis of complex molecules due to their ability to methylate or alkylate substrates with high regio-, chemo-, and stereoselectivity. Recent advancements in enzyme-catalyzed S-methylation and S-alkylation of S-adenosyl homocysteine (SAH) using synthetic alkylation agents have expanded the scope of methyltransferases in preparative biocatalysis. This development has transformed SAH from an unwanted byproduct into a crucial - and currently expensive - reagent. In this report, we present a simple and scalable one-pot synthesis of SAH, starting from racemic homocysteine thiolactone and adenosine. This process is catalyzed by recombinant α-amino-ε-caprolactam racemase, bleomycin hydrolase, and SAH hydrolase. The reaction proceeds to completion with near-stoichiometric mixtures of reactants, driven by the irreversible and stereoselective hydrolysis of thiolactone, followed by the thermodynamically favorable condensation of homocysteine with adenosine. We demonstrate that this method can be utilized to supplement preparative methylation reactions with SAH as a cofactor, as well as to synthesize and screen S-nucleosyl homocysteine derivatives in the search for stabilized SAM analogs.
| S-EPMC11391342 | biostudies-literature
Project description:Boolean satisfiability (k-SAT) is an NP-complete (k ≥ 3) problem that constitute one of the hardest classes of constraint satisfaction problems. In this work, we provide a proof of concept hardware based analog k-SAT solver, that is built using Magnetic Tunnel Junctions (MTJs). The inherent physics of MTJs, enhanced by device level modifications, is harnessed here to emulate the intricate dynamics of an analog satisfiability (SAT) solver. In the presence of thermal noise, the MTJ based system can successfully solve Boolean satisfiability problems. Most importantly, our results exhibit that, the proposed MTJ based hardware SAT solver is capable of finding a solution to a significant fraction (at least 85%) of hard 3-SAT problems, within a time that has a polynomial relationship with the number of variables(<50).
| S-EPMC5932068 | biostudies-literature
Project description:Lysine methylation has been extensively studied in histones, where it has been shown to provide specific epigenetic marks for the regulation of gene expression; however, the molecular mechanism and physiological function of lysine methylation in proteins other than histones remains to be fully addressed. To better understand the substrate diversity of lysine methylation, S-adenosylmethionine (SAM) derivatives with alkyne-moieties have been synthesized. A selenium-based SAM analog, propargylic Se-adenosyl-l-selenomethionine (ProSeAM), has a wide spectrum of reactivity against various lysine methyltransferases (KMTs) with sufficient stability to support enzymatic reactions in vitro. By using ProSeAM as a chemical probe for lysine methylation, we identified substrates for two seven-beta-strand KMTs, METTL21A and METTL10, on a proteomic scale in mammalian cells. METTL21A has been characterized as a heat shock protein (HSP)-70 methyltransferase. Mammalian METTL10 remains functionally uncharacterized, although its ortholog in yeast, See1, has been shown to methylate the translation elongation factor eEF1A. By using ProSeAM-mediated alkylation followed by purification and quantitative MS analysis, we confirmed that METTL21A labels HSP70 family proteins. Furthermore, we demonstrated that METTL10 also methylates the eukaryotic elongation factor EF1A1 in mammalian cells. Subsequent biochemical characterization revealed that METTL10 specifically trimethylates EF1A1 at lysine 318 and that siRNA-mediated knockdown of METTL10 decreases EF1A1 methylation levels in vivo. Thus, our study emphasizes the utility of the synthetic cofactor ProSeAM as a chemical probe for the identification of non-histone substrates of KMTs.
| S-EPMC4140779 | biostudies-literature