RESUMEN
Heavy metal pollution in roadside soil may harm humans, animals, plants, and local ecosystems. This study aimed to explore the sources and potential ecological risks of heavy metals in soils of roadside trees under different land uses, using soil samples collected from 136 roads across 16 administrative districts in Shanghai. The contents, pollution characteristics, potential ecological risks, and sources of seven heavy metals were analyzed, including Cr, Ni, Cd, Pb, As, Cu, and Zn. Results showed that (1) land use patterns affected the heavy metal contents, with industrial and construction areas showing higher contents while agricultural and forestry areas lower; (2) the ranking of heavy metal pollution levels was Cd > As > Pb > Cu > Ni > Cr > Zn. Cd exhibited the highest potential ecological risk, falling within the moderate to considerable potential ecological risk interval; (3) the sources of Cu, Zn, Cr, Ni, Cd, and Pb were associated with traffic emissions, whereas As had independent other sources and Pb in industrial and construction areas was also influenced by industrial emissions. These results provide valuable references on the control of heavy metal pollutants and the management of land uses in megacities.
Asunto(s)
Metales Pesados , Contaminantes del Suelo , Humanos , Suelo , Monitoreo del Ambiente , Árboles , Ecosistema , Cadmio , Plomo , Contaminantes del Suelo/análisis , China , Metales Pesados/análisis , Medición de RiesgoRESUMEN
Cyclic AMP receptor protein (CRP) is one of the seven global regulators in Escherichia coli, which regulates the expression of over 490 genes. It contains a cAMP binding N-terminal domain and a DNA binding C-terminal domain, connected via a short hinge region. Various stress-tolerant E. coli mutants had been obtained through transcriptional engineering of CRP. This review aims to shed some light on the possible mechanism behind these CRP variants, from the change in CRP structure, transcription profile, and DNA binding affinity. The amino acid substitutions are distributed along the protein-certain mutations have shown higher frequency than others, such as T127N and D138Y. ß-Galactosidase reporter gene assay revealed that CRP mutants had lower binding affinity with all three classes of CRP-dependent promoters as compared to native CRP, which probably would change cellular transcription profile. Different CRP mutants would induce different cellular transcription profile in E. coli, but there are common genes differentially expressed in these variants, including upregulated gadAB and downregulated nontransporter genes aspA and tnaA, and transporter/poringenes malE, mglB, cstA, and lamB. We believe that transcriptional engineering of CRP can provide an alternative strain engineering method for E. coli and its detailed mechanism may need further investigations.
Asunto(s)
Antibacterianos/farmacología , Biotecnología/métodos , Proteína Receptora de AMP Cíclico/metabolismo , Farmacorresistencia Bacteriana , Tolerancia a Medicamentos , Proteínas de Escherichia coli/metabolismo , Escherichia coli/efectos de los fármacos , Escherichia coli/fisiología , Proteína Receptora de AMP Cíclico/genética , Proteínas de Escherichia coli/genética , Regulación Bacteriana de la Expresión Génica , Ingeniería Metabólica/métodos , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Estrés Fisiológico , Transcripción GenéticaRESUMEN
Bioprocesses such as production of organic acids or acid hydrolysis of bioresources during biofuel production often suffer limitations due to microbial sensitivity under acidic conditions. Approaches for improving the acid tolerance of these microbes have mainly focused on using metabolic engineering tools. Here, we tried to improve strain acidic tolerance from its transcription level, i.e. we adopted error-prone PCR method to engineer global regulator cAMP receptor protein (CRP) of Escherichia coli to improve its performance at low pH. The best mutant AcM1 was identified from random mutagenesis libraries based on its growth performance. AcM1 almost doubled (0.113h(-1)) the growth rate of the control (0.062h(-1)) at pH 4.24. It also demonstrated better thermotolerance than the control at 48°C, whose growth was completely inhibited at this temperature. Quantitative real time reverse transcription PCR results revealed a stress response overlap among low pH stress-, oxidative stress- and osmotic stress-related genes. The chief enzyme responsible for cell acid tolerance, glutamate decarboxylase, demonstrated over twofold activity in AcM1 compared to the control. Differential binding properties of AcM1 mutant CRP with Class-I, II, and III CRP-dependent promoters suggested that modifications to native CRP may lead to transcription profile changes. Hence, we believe that transcriptional engineering of global regulator CRP can provide a new strain engineering alternative for E. coli.
Asunto(s)
Ácido Acético/metabolismo , Adaptación Fisiológica/genética , Proteína Receptora de AMP Cíclico/genética , Proteínas de Escherichia coli/genética , Escherichia coli/fisiología , Ácido Succínico/metabolismo , Sitios de Unión/genética , Proteína Receptora de AMP Cíclico/metabolismo , Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Regulación Bacteriana de la Expresión Génica , Genes Bacterianos , Glutamato Descarboxilasa/genética , Glutamato Descarboxilasa/metabolismo , Ingeniería Metabólica , Mutagénesis , Regiones Promotoras Genéticas , TemperaturaRESUMEN
The limited isobutanol tolerance of Escherichia coli is a major drawback during fermentative isobutanol production. Different from classical strain engineering approaches, this work was initiated to improve E. coli isobutanol tolerance from its transcriptional level by engineering its global transcription factor cAMP receptor protein (CRP). Random mutagenesis libraries were generated by error-prone PCR of crp, and the libraries were subjected to isobutanol stress for selection. Variant IB2 (S179P, H199R) was isolated and exhibited much better growth (0.18 h(-1) ) than the control (0.05 h(-1) ) in 1.2% (v/v) isobutanol (9.6 g/L). Genome-wide DNA microarray analysis revealed that 58 and 308 genes in IB2 had differential expression (>2-fold, p < 0.05) in the absence and presence of 1% (v/v) isobutanol, respectively. When challenged with isobutanol, genes related to acid resistance (gadABCE, hdeABD), nitrate reduction (narUZYWV), flagella and fimbrial activity (lfhA, yehB, ycgR, fimCDF), and sulfate reduction and transportation (cysIJH, cysC, cysN) were the major functional groups that were up-regulated, whereas most of the down-regulated genes were enzyme (tnaA) and transporters (proVWX, manXYZ). As demonstrated by single-gene knockout experiments, gadX, nirB, rhaS, hdeB, and ybaS were found associated with strain isobutanol resistance. The intracellular reactive oxygen species (ROS) level in IB2 was only half of that of the control when facing stress, indicating that IB2 can withstand toxic isobutanol much better than the control.