Climatic change is an increasing challenge for agriculture that is driving the development of suitable crops in order to ensure supply for both human nutrition and animal feed. In this context, it is increasingly important to understand the biochemical responses of cells to environmental cues at the whole system level, an aim that is being brought closer by advances in high throughput, cost-efficient plant metabolomics. To support molecular breeding activities, we have assessed the economic, technical and statistical feasibility of using direct mass spectrometry methods to evaluate the physiological state of maize (Zea mays L.) plants grown under different stress conditions.
Nanoparticles are a promising cancer therapy for their use as drug carriers given their versatile functionalization with polyethylene glycol and proteins that can be recognized by overexpressed receptors in tumor cells. However, it has been suggested that in biological fluids, proteins cover nanoparticles, which gives the proteins a biological identity that could be responsible for unexpected biological responses: the so-called protein corona. A relevant biological event that is usually ignored in protein-corona formation is the interspecies differences in protein binding, which can be involved in the discrepancies observed in preclinical studies and the nanoparticle safety and efficiency. Hence, the aim of this study was to determine the differences between human and mouse plasma protein corona profiles in an active therapy model using silicon dioxide nanoparticles (SiO nanoparticles) functionalized with polyethylene glycol and transferrin. Functionalized SiO nanoparticles were made with a primary particle size of 25 nm and a transferrin content of 50 μg mg of nanoparticles and were PEGylated with a cross-linker. The proteomic analysis by nanoliquid chromatography tandem-mass spectrometry (nanoLC-MS/MS) showed interspecies differences. The most abundant proteins found in the human protein corona profile were immunoglobulins, actin cytoplasmic 1, hemoglobin subunit beta, serotransferrin, ficolin-3, complement C3, and apolipoprotein A-1. Meanwhile, the mouse protein corona adsorbed the serine protease inhibitor A3K, serotransferrin, alpha-1-antitrypsin 1-2, hemoglobin subunit beta, and fibrinogen gamma and beta chains. These protein-corona profile differences in the functionalized SiO nanoparticles indicate that biological responses observed in in vivo models could not be translated to clinical use and must be considered in the interpretation of preclinical trials in order to design more efficient and safer nanomedicines.
High-throughput metabolic phenotyping is a challenge, but it provides an alternative and comprehensive access to the rapid and accurate characterization of plants. In addition to the technical issues of obtaining quantitative data of plenty of metabolic traits from numerous samples, a suitable data processing and statistical evaluation strategy must be developed. We present a simple, robust and highly scalable strategy for the comparison of multiple chemical profiles from coffee and tea leaf extracts, based on direct-injection electrospray mass spectrometry (DIESI-MS) and hierarchical cluster analysis (HCA). More than 3500 individual Coffea canephora and Coffea arabica trees from experimental fields in Mexico were sampled and processed using this method. Our strategy permits the classification of trees according to their metabolic fingerprints and the screening for families with desired characteristics, such as extraordinarily high or low caffeine content in their leaves.
Mitochondrial porins or voltage-dependent anion channels (VDAC) are the main route for solute transport through outer mitochondrial membranes (OMM). In mammals, hexokinase (HK) binds to VDAC, which allows the channeling of ATP synthesized by oxidative phosphorylation toward HK. In plants, although HK has been found associated with OMM, evidence for an interaction with VDAC is scarce. Thus, in this work, we studied the physical and functional interaction between these proteins in beetroot mitochondria. To observe a physical interaction between HK and VDAC, OMM presenting HK activity were prepared from purified mitochondria. Protein complexes were solubilized from OMM with mild detergents and separated by centrifugation in glycerol gradients. Both HK activity and immunodetected VDAC were found in small (9S-13S) and large (>40S) complexes. OMM proteins were also separated according to their hydropathy by serial phase partitioning with Triton X-114. Most of HK activity was found in hydrophobic fractions where VDAC was also present. These results indicated that HK could be bound to VDAC in beetroot mitochondria. The functional interaction of HK with VDAC was demonstrated by observing the effect of apyrase on HK-catalyzed glucose phosphorylation in intact mitochondria. Apyrase, which hydrolyzes freely soluble ATP, competed efficiently with hexokinase for ATP when it was produced outside mitochondria (with PEP and pyruvate kinase), but not when it was produced inside mitochondria by oxidative phosphorylation. These results suggest that HK closely interacts with VDAC in beetroot mitochondria, and that this interaction allows the channeling of respiratory ATP toward HK through VDAC.
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