The specific radioactivity of the precursor pool for estimates of the rate of protein synthesis.
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ABSTRACT:
SUBMITTER: Fern EB
PROVIDER: S-EPMC1177923 | biostudies-other | 1973 Aug
REPOSITORIES: biostudies-other
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Project description:1. Rats were infused in vivo with [U-(14)C]glycine for periods of 2-6h, during which time the specific radioactivity of the free glycine in plasma and tissue approached a constant value. 2. Free serine also became labelled. The ratio of specific radioactivity of serine to that of glycine in the protein of liver, kidney, brain, jejunum, heart, diaphragm and gastrocnemius muscle was closer to the ratio in the free amino acid pool of the tissue than that of the plasma. 3. The kinetics of incorporation of [(14)C]glycine and [(14)C]serine into the protein of gastrocnemius muscle further suggested that the plasma free amino acids were not the immediate precursors of protein. 4. Infusion of rats with [U-(14)C]serine resulted in labelling of free glycine. The ratio of specific radioactivity of glycine to serine in the protein of liver, kidney, brain, jejunum and heart again suggested incorporation from a pool similar to the free amino acid pool of the tissue. 5. Rates of tissue protein synthesis calculated from the incorporation into protein of both radioactive glycine and serine, either infused or derived, were very similar when the precursor specific radioactivity was taken to be that in the total free amino acids of the tissue. Except for gastrocnemius muscle and diaphragm during the infusion of radioactive serine, the rates of tissue protein synthesis calculated from the specific radioactivity of the free glycine and serine in plasma differed markedly.
| S-EPMC1168293 | biostudies-other
Project description:All organisms on Earth are exposed to low doses of natural radioactivity but some habitats are more radioactive than others. Yet, documenting the influence of natural radioactivity on the evolution of biodiversity is challenging. Here, we addressed whether organisms living in naturally more radioactive habitats accumulate more mutations across generations using 14 species of waterlice living in subterranean habitats with contrasted levels of radioactivity. We found that the mitochondrial and nuclear mutation rates across a waterlouse species' genome increased on average by 60% and 30%, respectively, when radioactivity increased by a factor of three. We also found a positive correlation between the level of radioactivity and the probability of G to T (and complementary C to A) mutations, a hallmark of oxidative stress. We conclude that even low doses of natural bedrock radioactivity influence the mutation rate possibly through the accumulation of oxidative damage, in particular in the mitochondrial genome.
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Project description:The relative molar synthesis of cardiac contractile proteins has been measured in the perfused heart under control haemodynamic conditions. This synthesis, of myosin heavy chains, individual light chains (1 and 2), actin and tropomyosin, was determined from isolated guinea-pig hearts perfused for 3h simultaneously with constant specific radioactivities and concentrations of [3H]lysine and [3H]phenylalanine. The data strongly suggest that all of the proteins studied were synthesized from the same precursor pools of lysine and phenylalanine, since the ratio of the specific activities of the two labels was the same in all of the proteins. Measurement of molar synthesis of each contractile protein was the same with either labelled amino acid. Under control haemodynamic-perfusion conditions, the relative molar synthesis of the contractile proteins was actin greater than heavy chains greater than light chain 2 greater than light chain 1 greater than tropomyosin.
| S-EPMC1162800 | biostudies-other
Project description:Human [[3,5-3H2]Tyr23]corticotropin-(1-39)-nonatriacontapeptide of specific radioactivity 25.2 Ci/mmol, identical with unlabelled human corticotropin by several criteria, was prepared via the fully protected di-iodotyrosine compound. The latter was synthesized by classical procedures.
| S-EPMC1164883 | biostudies-other
Project description:The availability of several bioorthogonal reactions that can proceed selectively and efficiently under physiologically relevant conditions has garnered the interest of biochemists and organic chemists alike. Bioorthogonal cleavage reactions represent the latest innovation in click chemistry. Here, we employed the Staudinger ligation reaction to release radioactivity from immunoconjugates, improving target-to-background ratios. In this proof-of-concept study, model systems, including the anti-HER2 antibody trastuzumab, radioisotope I-131, and a newly synthesized bifunctional phosphine, were used. Staudinger ligation occurred when biocompatible N-glycosyl azides reacted with this radiolabeled immunoconjugate, leading to cleavage of the radioactive label from the molecule. We demonstrated this click cleavage in vitro and in vivo. Biodistribution studies in tumor models showed that radioactivity was eliminated from the bloodstream, thereby improving tumor-to-blood ratios. SPECT imaging revealed that tumors could be visualized with enhanced clarity. Our simple approach represents a novel application of bioorthogonal click chemistry in the development of antibody-based theranostics.
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Project description:Evidence is presented that in human granulocytes the immediate precursor pool of phenylalanine for protein synthesis in vitro is intracellular significant compartmentation.
| S-EPMC1172526 | biostudies-other
Project description:PurposeTo assess the accuracy of morphing an established reference electromagnetic head model to a subject-specific morphometry for the estimation of specific absorption rate (SAR) in 7T parallel-transmit (pTx) MRI.MethodsSynthetic T1 -weighted MR images were created from three high-resolution open-source electromagnetic head voxel models. The accuracy of morphing a "reference" (multimodal image-based detailed anatomical [MIDA]) electromagnetic model into a different subject's native space (Duke and Ella) was compared. Both linear and nonlinear registration methods were evaluated. Maximum 10-g averaged SAR was estimated for circularly polarized mode and for 5000 random RF shim sets in an eight-channel transmit head coil, and comparison made between the morphed MIDA electromagnetic models and the native Duke and Ella electromagnetic models, respectively.ResultsThe averaged error in maximum 10-g averaged SAR estimation across pTx MRI shim sets between the MIDA and the Duke target model was reduced from 17.5% with only rigid-body registration, to 11.8% when affine linear registration was used, and further reduced to 10.7% when nonlinear registration was used. The corresponding figures for the Ella model were 16.7%, 11.2%, and 10.1%.ConclusionWe found that morphometry accounts for up to half of the subject-specific differences in pTx SAR. Both linear and nonlinear morphing of an electromagnetic model into a target subject improved SAR agreement by better matching head size, morphometry, and position. However, differences remained, likely arising from details in tissue composition estimation. Thus, the uncertainty of the head morphometry and tissue composition may need to be considered separately to achieve personalized SAR estimation.
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Project description:Secreted amyloid precursor protein-? (sAPP?) is a neuroprotective and memory-enhancing molecule, however, the mechanisms through which sAPP? promotes these effects are not well understood. Recently, we have shown that sAPP? enhances cell-surface expression of glutamate receptors. Activity-related cytoskeletal-associated protein Arc (Arg3.1) is an immediate early gene capable of modulating long-term potentiation, long-term depression and homeostatic plasticity through regulation of ?-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor localization. Accordingly, we hypothesized that sAPP? may enhance synaptic plasticity, in part, by the de novo synthesis of Arc. Using primary cortical and hippocampal neuronal cultures we found that sAPP? (1 nM, 2 h) enhances levels of Arc mRNA and protein. Arc protein levels were increased in both the neuronal somata and dendrites in a Ca2+/calmodulin-dependent protein kinase II-dependent manner. Additionally, dendritic Arc expression was dependent upon activation of mitogen-activated protein kinase and protein kinase G. The enhancement of dendritic Arc protein was significantly reduced by antagonism of N-methyl-D-aspartate (NMDA) and nicotinic acetylcholine (?7nACh) receptors, and fully eliminated by dual application of these antagonists. This effect was further corroborated in area CA1 of acute hippocampal slices. These data suggest sAPP?-regulated plasticity within hippocampal neurons is mediated by cooperation of NMDA and ?7nACh receptors to engage a cascade of signal transduction molecules to enhance the transcription and translation of Arc.
| S-EPMC6702288 | biostudies-literature
Project description:Hypoxic non-small cell lung cancer (NSCLC) is dependent on Notch-1 signaling for survival. Targeting Notch-1 by means of γ-secretase inhibitors (GSI) proved effective in killing hypoxic NSCLC. Post-mortem analysis of GSI-treated, NSCLC-burdened mice suggested enhanced phosphorylation of 4E-BP1 at threonines 37/46 in hypoxic tumor tissues. In vitro dissection of this phenomenon revealed that Amyloid Precursor Protein (APP) inhibition was responsible for a non-canonical 4E-BP1 phosphorylation pattern rearrangement-a process, in part, mediated by APP regulation of the pseudophosphatase Styx. Upon APP depletion we observed modifications of eIF-4F composition indicating increased recruitment of eIF-4A to the mRNA cap. This phenomenon was supported by the observation that cells with depleted APP were partially resistant to silvestrol, an antibiotic that interferes with eIF-4A assembly into eIF-4F complexes. APP downregulation in dividing human cells increased the rate of global protein synthesis, both cap- and IRES-dependent. Such an increase seemed independent of mTOR inhibition. After administration of Torin-1, APP downregulation and Mechanistic Target of Rapamycin Complex 1 (mTORC-1) inhibition affected 4E-BP1 phosphorylation and global protein synthesis in opposite fashions. Additional investigations indicated that APP operates independently of mTORC-1. Key phenomena described in this study were reversed by overexpression of the APP C-terminal domain. The presented data suggest that APP may be a novel regulator of protein synthesis in dividing human cells, both cancerous and primary. Furthermore, APP appears to affect translation initiation using mechanisms seemingly dissimilar to mTORC-1 regulation of cap-dependent protein synthesis.
| S-EPMC4445069 | biostudies-literature
Project description:Previous studies from this laboratory on protein turnover in 3H-labelled L-cell cultures have shown recovery of total 3H at the end of a 3-day experiment to be always significantly in excess of the 3H recovered at the beginning of the experiment. In this study we have critically reviewed a number of possible sources for this error in measuring radioactivity in cell proteins. 3H-labelled proteins, when dissolved in 0.3 M-NaOH and counted for radioactivity in a liquid-scintillation spectrometer, showed losses of 30-40% of the radioactivity; neither external or internal standardization compensated for this loss. Hydrolysis of these proteins with either Pronase or concentrated HCl significantly increased the measured radioactivity. In addition, approx. 5-10% of the cell protein is left on the plastic culture dish when cells are recovered in phosphate-buffered saline. To aggravate this latter loss further, this surface-adherent protein, after pulse labelling, contains proteins of high radioactivity that turn over rapidly and make a major contribution to the accumulating radioactivity in the medium. These combined errors can account for up to 60% of the total radioactivity in the cell culture. Similar analytical errors have been found in studies of other cell cultures. The effect of these analytical errors on estimates of protein turnover in cell cultures is discussed.
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