RESUMEN
Galectins are a large and diverse protein family defined by the presence of a carbohydrate recognition domain (CRD) that binds ß-galactosides. They play important roles in early development, tissue regeneration, immune homeostasis, pathogen recognition, and cancer. In many cases, studies that examine galectin biology and the effect of manipulating galectins are aided by, or require the ability to express and purify, specific members of the galectin family. In many cases, E. coli is employed as a heterologous expression system, and galectin expression is induced with isopropyl ß-galactoside (IPTG). Here, we show that galectin-3 recognizes IPTG with micromolar affinity and that as IPTG induces expression, newly synthesized galectin can bind and sequester cytosolic IPTG, potentially repressing further expression. To circumvent this putative inhibitory feedback loop, we utilized an autoinduction protocol that lacks IPTG, leading to significantly increased yields of galectin-3. Much of this work was done within the context of a course-based undergraduate research experience, indicating the ease and reproducibility of the resulting expression and purification protocols.
Asunto(s)
Escherichia coli , Galectina 3 , Isopropil Tiogalactósido , Galectina 3/genética , Galectina 3/metabolismo , Galectina 3/biosíntesis , Galectina 3/química , Escherichia coli/genética , Escherichia coli/metabolismo , Humanos , Isopropil Tiogalactósido/farmacología , Expresión Génica , Galectinas/genética , Galectinas/metabolismo , Galectinas/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/aislamiento & purificación , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/química , Proteínas Sanguíneas/genética , Proteínas Sanguíneas/metabolismoRESUMEN
Crop yields are sensitive to extreme weather events. Improving the understanding of the mechanisms and the drivers of the projection uncertainties can help to improve decisions. Previous studies have provided important insights, but often sample only a small subset of potentially important uncertainties. Here we expand on a previous statistical modeling approach by refining the analyses of two uncertainty sources. Specifically, we assess the effects of uncertainties surrounding crop-yield model parameters and climate forcings on projected crop yield. We focus on maize yield projections in the eastern U.S.in this century. We quantify how considering more uncertainties expands the lower tail of yield projections. We characterized the relative importance of each uncertainty source and show that the uncertainty surrounding yield model parameters is the main driver of yield projection uncertainty.
Asunto(s)
Zea mays/crecimiento & desarrollo , Producción de Cultivos , Modelos Estadísticos , Incertidumbre , Estados UnidosRESUMEN
Crude oil contaminated soil cores were collected from a basin that contained oily solids left from three decades of oil production. Hydrocarbon biomarker analyses revealed that the soil extracts were moderately biodegraded compared with the non-degraded source oil. The degree of biodegradation also decreased with core depth (7 cm to 1 m). These data were correlated to compositional changes observed in acidic NSO-compounds that were selectively ionized and mass resolved by negative ion electrospray Fourier transform ion cyclotron resonance mass spectrometry (ESI FT-ICR MS). Among the NSO-compounds ionized, the increase in naphthenic acid concentration (e.g., acyclic and alicyclic carboxylic acids) best correlated with the increase in biodegradation (e.g., from non-degraded to moderately degraded) as determined by the hydrocarbon biomarker analyses. The most biodegraded surface extracts (7 cm) exhibited an 80% increase in the abundance of acids relative to the source oil. Use of an internal standard allowed the semi-quantitative determination of the total naphthenic acid concentration, which decreased significantly (P < 0.05) with soil depth. Furthermore, the shift to higher double bond equivalents (DBEs), from acyclic to alicyclic acids, indicated that the increase in acids in the soil extracts was predominantly due to biotic processes. This work demonstrates the potential of ESI FT-ICR MS as a semi-quantitative tool to monitor the production of naphthenic acids during crude oil biotransformation in the environment.