RESUMO
In this review the current state-of-the-art of S-adenosylmethionine (SAM)-dependent methyltransferases and SAM are evaluated. Their structural classification and diversity is introduced and key mechanistic aspects presented which are then detailed further. Then, catalytic SAM as a target for drugs, and approaches to utilise SAM as a cofactor in synthesis are introduced with different supply and regeneration approaches evaluated. The use of SAM analogues are also described. Finally O-, N-, C- and S-MTs, their synthetic applications and potential for compound diversification is given.
Assuntos
Metiltransferases , S-Adenosilmetionina , Metiltransferases/química , S-Adenosilmetionina/químicaRESUMO
S-Adenosyl-L-homocysteine hydrolase (SAHH) reversibly cleaves S-adenosyl-L-homocysteine, the product of S-adenosyl-L-methionine-dependent methylation reactions. The conversion of S-adenosyl-L-homocysteine into adenosine and L-homocysteine plays an important role in the regulation of the methyl cycle. An alternative metabolic route for S-adenosyl-L-methionine regeneration in the extremophiles Methanocaldococcus jannaschii and Thermotoga maritima has been identified, featuring the deamination of S-adenosyl-L-homocysteine to S-inosyl-L-homocysteine. Herein, we report the structural characterisation of different archaeal SAHHs together with a biochemical analysis of various SAHHs from all three domains of life. Homologues deriving from the Euryarchaeota phylum show a higher conversion rate with S-inosyl-L-homocysteine compared to S-adenosyl-L-homocysteine. Crystal structures of SAHH originating from Pyrococcus furiosus in complex with SLH and inosine as ligands, show architectural flexibility in the active site and offer deeper insights into the binding mode of hypoxanthine-containing substrates. Altogether, the findings of our study support the understanding of an alternative metabolic route for S-adenosyl-L-methionine and offer insights into the evolutionary progression and diversification of SAHHs involved in methyl and purine salvage pathways.
Assuntos
Archaea , S-Adenosilmetionina , S-Adenosilmetionina/química , S-Adenosilmetionina/metabolismo , Archaea/metabolismo , Adenosina/metabolismo , Metionina , HomocisteínaAssuntos
Poluentes Atmosféricos/toxicidade , Biomassa , Brônquios/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Perfilação da Expressão Gênica/métodos , Material Particulado/toxicidade , Madeira , Brônquios/metabolismo , Linhagem Celular , Células Epiteliais/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Redes Reguladoras de Genes/efeitos dos fármacos , Humanos , Lipopolissacarídeos/toxicidade , Análise de Sequência com Séries de Oligonucleotídeos , Tamanho da Partícula , Centrais Elétricas , RNA de Cadeia Dupla/toxicidade , Medição de Risco , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Transcriptoma/efeitos dos fármacosRESUMO
The ubiquitous cofactor S-adenosyl-l-methionine (SAM) is part of numerous biochemical reactions in metabolism, epigenetics, and cancer development. As methylation usually improves physiochemical properties of compounds relevant for pharmaceutical use, the sustainable use of SAM as a methyl donor in biotechnological applications is an important goal. SAM-dependent methyltransferases are consequently an emerging biocatalytic tool for environmentally friendly and selective alkylations. However, SAM shows undesirable characteristics such as degradation under mild conditions and its stoichiometric use is economically not reasonable. Here, we report an optimised biomimetic system for the regeneration of SAM and SAM analogues consisting of effective nucleoside triphosphate formation and an additional l-methionine regeneration cycle without by-product accumulation. The bicyclic system uses seven enzymes, S-methylmethionine as methyl donor and a surplus of inorganic polyphosphate, along with catalytic amounts of l-methionine and cofactor building block reaching conversions of up to 99% (up to 200 turnovers). We also show that the cycle can be run with cofactor building blocks containing different purine and pyrimidine nucleobases, which can be fed in at the nucleoside or nucleotide stage. These alternative cofactors are in turn converted to the corresponding SAM analogues, which are considered to be a key for the development of bioorthogonal systems. In addition to purified enzymes, the bicyclic system can also be used with crude lysates highlighting its broad biocatalytic applicability.
RESUMO
Nearly half of the world's population relies on combustion of solid biofuels to cover fundamental energy demands. Epidemiologic data demonstrate that particularly long-term emissions adversely affect human health. However, pathological molecular mechanisms are insufficiently characterized. Here we demonstrate that long-term exposure to fine particulate matter (PM2.5) from biomass combustion had no impact on cellular viability and proliferation but increased intracellular reactive oxygen species (ROS) levels in bronchial epithelial BEAS-2B cells. Exposure to PM2.5 induced the nuclear factor erythroid 2-related factor 2 (Nrf2) and mediated an anti-oxidative response, including enhanced levels of intracellular glutathione (GSH) and nuclear accumulation of heme oxygenase-1 (HO-1). Activation of Nrf2 was promoted by the c-Jun N-terminal kinase JNK1/2, but not p38 or Akt, which were also induced by PM2.5. Furthermore, cells exposed to PM2.5 acquired chemoresistance to doxorubicin, which was associated with inhibition of apoptosis and elevated levels of GSH in these cells. Our findings propose that exposure to PM2.5 induces molecular defense mechanisms, which prevent cellular damage and may thus explain the initially relative rare complications associated with PM2.5. However, consistent induction of pro-survival pathways may also promote the progression of diseases. Environmental conditions inducing anti-oxidative responses may have the potential to promote a chemoresistant cellular phenotype.
Assuntos
Resistencia a Medicamentos Antineoplásicos , Neoplasias , Material Particulado/toxicidade , Biomassa , Células Epiteliais , Humanos , Estresse Oxidativo , Material Particulado/análise , Espécies Reativas de OxigênioRESUMO
Metalloproteinase inhibitors often feature hydroxamate moieties to facilitate the chelation of metal ions in the catalytic center of target enzymes. Actinonin and matlystatins are potent metalloproteinase inhibitors that comprise rare N-hydroxy-2-pentyl-succinamic acid warheads. Here we report the identification and characterization of their biosynthetic pathways. By gene cluster comparison and a combination of precursor feeding studies, heterologous pathway expression and gene deletion experiments we are able to show that the N-hydroxy-alkyl-succinamic acid warhead is generated by an unprecedented variation of the ethylmalonyl-CoA pathway. Moreover, we present evidence that the remarkable structural diversity of matlystatin congeners originates from the activity of a decarboxylase-dehydrogenase enzyme with high similarity to enzymes that form epoxyketones. We further exploit this mechanism to direct the biosynthesis of non-natural matlystatin derivatives. Our work paves the way for follow-up studies on these fascinating pathways and allows the identification of new protease inhibitors by genome mining.