scholarly journals A conformational change in the N terminus of SLC38A9 signals mTORC1 activation

2018 ◽  
Author(s):  
Ma Jinming ◽  
Hsiang-Ting Lei ◽  
Tamir Gonen
Keyword(s):  
Amino Acids ◽  
Conformational Change ◽  
Energy Levels ◽  
Binding Pocket ◽  
Chain Model ◽  
Fed State ◽  
Terminal Domain ◽  
Cellular Stresses ◽  
Significant Conformational Change ◽  
Mtorc1 Activation

AbstractmTORC1 is a central signal hub that integrates multiple environmental cues, such as cellular stresses, energy levels, nutrients and certain amino acids, to modulate metabolic status and cellular responses. Recently, SLC38A9, a lysosomal amino acid transporter, has emerged as a sensor for luminal arginine levels and as an activator of mTOCRC1. The activation of mTORC1 occurs through the N-terminal domain of SLC38A9. Here, we determined the crystal structure of SLC38A9 and surprisingly found its N-terminal fragment inserted deep into the transporter, bound in the substrate binding pocket where normally arginine would bind. Compared with our recent arginine bound structure of SLC38A9, a significant conformational change of the N-terminal domain was observed. A ball-and-chain model is proposed for mTORC1 activation where in the starved state the N-terminal domain of SLC38A9 is buried deep in the transporter but in the fed state the N-terminal domain could be released becoming free to bind the Rag GTPase complex and to activate mTORC1. This work provides important new insights into how SLC38A9 senses the fed state and activates the mTORC1 pathways in response to dietary amino acids.One Sentence SummaryN-plug inserted state of SLC38A9 reveals mechanisms of mTORC1 activation and arginine-enhanced luminal amino acids efflux.

LEVELS OF FREE PLASMA AMINO ACIDS UNDER ACUTE NORMOBARIC HYPOXIA IN VOLUNTEERS IN FASTED AND POSTPRANDIAL STATES

2020 ◽  
pp. 108-117
Author(s):  
A.A. Chernykh ◽  
N.N. Potolitsyna ◽  
E.A. Burykh ◽  
E.R. Boyko
Keyword(s):  
Amino Acids ◽  
Normobaric Hypoxia ◽  
Adaptation To Hypoxia ◽  
Plasma Amino Acids ◽  
Fed State ◽  
Metabolic Substrates ◽  
Overnight Fasting ◽  
Group 2 ◽  
Free Plasma ◽  
Group 1

The aim of the study was to assess the effect of acute normobaric hypoxia on free plasma amino acids (AA) in volunteers after overnight fasting and in the fed state. Materials and Methods. Group 1 (n=13, aged 22–32) participated in the study in the morning after overnight fasting. Group 2 (n=9, aged 22–32) took part in the study after a light fat-free breakfast. Acute normobaric hypoxia was achieved by breathing a hypoxic gas mixture (9 % O2 and 91 % N2) through a mask. According to the experimental protocol, blood sampling from the cubital vein was performed for analysis. Free plasma amino acids were analyzed using the Aracus amino acid analyzer. Results. Prior to the hypoxia onset, at the 5th and 20th minutes of hypoxia, no statistically significant differences in free AA levels were observed in the groups (p>0.05). At the 10th minute of hypoxia the levels of four AAs (serine, threonine, glutamine, and histidine) were significantly higher in Group 1 than in Group 2 (p<0.05). This was probably due to differences in functioning of several key “harmonizing” AA transporters (ASCT1 (SLC1A4), ASCT2 (SLC1A5) and LAT1 (SC7A5)), for which the AAs were metabolic substrates. It can be assumed, that such changes were caused by currently unclear mechanisms of fast regulation of AA transporter activity, associated with nutritional status. Conclusion. We believe that our findings may be important for providing better adaptation to hypoxia, and for more efficient correction of hypoxic negative effects. Keywords: acute normobaric hypoxia, free plasma amino acids, human. Цель исследования: изучить воздействие острой нормобарической гипоксии на метаболизм свободных аминокислот (АК) плазмы крови у добровольцев, участвовавших в исследовании натощак и после лёгкого завтрака. Материалы и методы. Первая группа добровольцев (22–32 года, n=13) участвовала в исследовании утром натощак, вторая группа (22–32 года, n=9) – через 2–3 ч после лёгкого безжирового завтрака. Гипоксия создавалась путём подачи через маску дыхательной смеси, содержащей 9 % О2 и 91 % N2. В соответствии с протоколом проводился периодический забор крови из локтевой вены для анализа. Оценка уровней свободных АК плазмы крови производилась с помощью аминокислотного анализатора Aracus. Результаты. До начала гипоксии, на 5-й и 20-й мин гипоксии уровни свободных АК в первой и второй группах значимо не различались (p>0,05). На 10-й мин гипоксии между первой и второй группами наблюдались статистически значимые различия уровней четырёх АК: глутамина, серина, треонина и гистидина (p<0,05). Это, вероятно, было обусловлено изменениями в работе «гармонизирующих» мембранных транспортёров (ASCT1 (SLC1A4), ASCT2 (SLC1A5) и LAT1 (SC7A5)), для которых эти АК являются обменными субстратами. Можно предположить, что данные изменения были опосредованы пока неясными механизмами быстрой регуляции активности этих транспортёров, зависящими от питания. Выводы. Мы полагаем, что полученные результаты могут иметь значение для обеспечения адаптации организма человека к острой гипоксии и эффективной коррекции последствий гипоксического воздействия. Ключевые слова: острая нормобарическая гипоксия, свободные аминокислоты плазмы крови, человек.

scholarly journals Determinants of the Endosomal Localization of Sorting Nexin 1

2005 ◽  
Vol 16 (4) ◽  
pp. 2049-2057 ◽  
Author(s):  
Qi Zhong ◽  
Martin J. Watson ◽  
Cheri S. Lazar ◽  
Andrea M. Hounslow ◽  
Jonathan P. Waltho ◽  
...  
Keyword(s):  
Binding Pocket ◽  
Early Endosome ◽  
Nmr Structure ◽  
Sorting Nexin ◽  
High Affinity ◽  
Terminal Domain ◽  
Px Domain ◽  
Phox Homology ◽  
Phosphatidylinositol 3

The sorting nexin (SNX) family of proteins is characterized by sequence-related phox homology (PX) domains. A minority of PX domains bind with high affinity to phosphatidylinositol 3-phosphate [PI(3)P], whereas the majority of PX domains exhibit low affinity that is insufficient to target them to vesicles. SNX1 is located on endosomes, but its low affinity PX domain fails to localize in vivo. The NMR structure of the PX domain of SNX1 reveals an overall fold that is similar to high-affinity PX domains. However, the phosphatidylinositol (PI) binding pocket of the SNX1 PX domain is incomplete; regions of the pocket that are well defined in high-affinity PX domains are highly mobile in SNX1. Some of this mobility is lost upon binding PI(3)P. The C-terminal domain of SNX1 is a long helical dimer that localizes to vesicles but not to the early endosome antigen-1–containing vesicles where endogenous SNX1 resides. Thus, the obligate dimerization of SNX1 that is driven by the C-terminal domain creates a high-affinity PI binding species that properly targets the holo protein to endosomes.

Negatively-charged amino acids at the peptide-binding pocket of HLA-DPB1 alleles are associated with susceptibility to anti-topoisomerase I-positive systemic sclerosis

2016 ◽  
Vol 77 (7) ◽  
pp. 550-554 ◽  
Author(s):  
Jeong Seok Lee ◽  
Jin Kyun Park ◽  
Heung Jae Kim ◽  
Hyung Ki Lee ◽  
Yeong Wook Song ◽  
...  
Keyword(s):  
Amino Acids ◽  
Systemic Sclerosis ◽  
Topoisomerase I ◽  
Peptide Binding ◽  
Binding Pocket ◽  
Charged Amino Acids

scholarly journals Insights into genome recoding from the mechanism of a classic +1-frameshifting tRNA

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Howard Gamper ◽  
Haixing Li ◽  
Isao Masuda ◽  
D. Miklos Robkis ◽  
Thomas Christian ◽  
...  
Keyword(s):  
Amino Acids ◽  
Conformational Change ◽  
Molecular Mechanism ◽  
Late Stage ◽  
Anticodon Loop ◽  
Pairing Mechanism ◽  
Extra Nucleotide ◽  
In Cells

AbstractWhile genome recoding using quadruplet codons to incorporate non-proteinogenic amino acids is attractive for biotechnology and bioengineering purposes, the mechanism through which such codons are translated is poorly understood. Here we investigate translation of quadruplet codons by a +1-frameshifting tRNA, SufB2, that contains an extra nucleotide in its anticodon loop. Natural post-transcriptional modification of SufB2 in cells prevents it from frameshifting using a quadruplet-pairing mechanism such that it preferentially employs a triplet-slippage mechanism. We show that SufB2 uses triplet anticodon-codon pairing in the 0-frame to initially decode the quadruplet codon, but subsequently shifts to the +1-frame during tRNA-mRNA translocation. SufB2 frameshifting involves perturbation of an essential ribosome conformational change that facilitates tRNA-mRNA movements at a late stage of the translocation reaction. Our results provide a molecular mechanism for SufB2-induced +1 frameshifting and suggest that engineering of a specific ribosome conformational change can improve the efficiency of genome recoding.

scholarly journals A role for BDNF- and NMDAR-induced lysosomal recruitment of mTORC1 in the regulation of neuronal mTORC1 activity

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Dany Khamsing ◽  
Solène Lebrun ◽  
Isabelle Fanget ◽  
Nathanaël Larochette ◽  
Christophe Tourain ◽  
...  
Keyword(s):  
Amino Acids ◽  
Nmda Receptors ◽  
Hippocampal Neurons ◽  
De Novo ◽  
Long Term Potentiation ◽  
Activation Mechanism ◽  
Functional Link ◽  
Term Potentiation ◽  
Endocytic Compartments ◽  
Mtorc1 Activation

AbstractMemory and long term potentiation require de novo protein synthesis. A key regulator of this process is mTORC1, a complex comprising the mTOR kinase. Growth factors activate mTORC1 via a pathway involving PI3-kinase, Akt, the TSC complex and the GTPase Rheb. In non-neuronal cells, translocation of mTORC1 to late endocytic compartments (LEs), where Rheb is enriched, is triggered by amino acids. However, the regulation of mTORC1 in neurons remains unclear. In mouse hippocampal neurons, we observed that BDNF and treatments activating NMDA receptors trigger a robust increase in mTORC1 activity. NMDA receptors activation induced a significant recruitment of mTOR onto lysosomes even in the absence of external amino acids, whereas mTORC1 was evenly distributed in neurons under resting conditions. NMDA receptor-induced mTOR translocation to LEs was partly dependent on the BDNF receptor TrkB, suggesting that BDNF contributes to the effect of NMDA receptors on mTORC1 translocation. In addition, the combination of Rheb overexpression and artificial mTORC1 targeting to LEs by means of a modified component of mTORC1 fused with a LE-targeting motif strongly activated mTOR. To gain spatial and temporal control over mTOR localization, we designed an optogenetic module based on light-sensitive dimerizers able to recruit mTOR on LEs. In cells expressing this optogenetic tool, mTOR was translocated to LEs upon photoactivation. In the absence of growth factor, this was not sufficient to activate mTORC1. In contrast, mTORC1 was potently activated by a combination of BDNF and photoactivation. The data demonstrate that two important triggers of synaptic plasticity, BDNF and NMDA receptors, synergistically power the two arms of the mTORC1 activation mechanism, i.e., mTORC1 translocation to LEs and Rheb activation. Moreover, they unmask a functional link between NMDA receptors and mTORC1 that could underlie the changes in the synaptic proteome associated with long-lasting changes in synaptic strength.

In silico study of binding affinity of nitrogenous bicyclic heterocycles: fragment-to-fragment approach

2020 ◽  
Vol 15 (2) ◽  
pp. 49-59
Author(s):  
Yevheniia Velihina ◽  
Nataliya Obernikhina ◽  
Stepan Pilyo ◽  
Maryna Kachaeva ◽  
Oleksiy Kachkovsky ◽  
...  
Keyword(s):  
Amino Acids ◽  
Binding Affinity ◽  
In Silico ◽  
Density Functional ◽  
Energy Levels ◽  
Aromatic Amino Acids ◽  
Electron System ◽  
High Energy ◽  
Proton Donor ◽  
Stabilization Energy

The binding affinity of model aromatic amino acids and heterocycles and their derivatives condensed with pyridine were investigated in silico and are presented in the framework of fragment-to-fragment approach. The presented model describes interaction between pharmacophores and biomolecules. Scrupulous data analysis shows that expansion of the π-electron system by heterocycles annelation causes the shifting up of high energy levels, while the appearance of new the dicoordinated nitrogen atom is accompanied by decreasing of the donor-acceptor properties. Density Functional Theory (DFT) wB97XD/6-31(d,p)/calculations of π-complexes of the heterocycles 1-3 with model fragments of aromatic amino acids, which were formed by π-stack interaction, show an increase in the stabilization energy of π-complexes during the moving from phenylalanine to tryptophan. DFT calculation of pharmacophore complexes with model proton-donor amino acid by the hydrogen bonding mechanism (H-B complex) shows that stabilization energy (DE) increases from monoheterocycles to their condensed derivatives. The expansion of the π-electron system by introducing phenyl radicals to the oxazole cycle as reported earlier [18] leads to a decrease in the stabilization energy of the [Pharm-BioM] complexes in comparison with the annelated oxazole by the pyridine cycle.

scholarly journals Role of a tryptophan anchor in human topoisomerase I structure, function and inhibition

2008 ◽  
Vol 411 (3) ◽  
pp. 523-530 ◽  
Author(s):  
Gary S. Laco ◽  
Yves Pommier
Keyword(s):  
Amino Acids ◽  
Topoisomerase I ◽  
Electrostatic Interactions ◽  
Functional Domain ◽  
Site Mutation ◽  
Supercoiled Dna ◽  
Terminal Domain ◽  
Tryptophan Residues ◽  
N Terminus

Human Top1 (topoisomerase I) relaxes supercoiled DNA during cell division and transcription. Top1 is composed of 765 amino acids and contains an unstructured N-terminal domain of 200 amino acids, and a structured functional domain of 565 amino acids that binds and relaxes supercoiled DNA. In the present study we examined the region spanning the junction of the N-terminal domain and functional domain (junction region). Analysis of several published Top1 structures revealed that three tryptophan residues formed a network of aromatic stacking interactions and electrostatic interactions that anchored the N-terminus of the functional domain to sub-domains containing the nose cone and active site. Mutation of the three tryptophan residues (Trp203/Trp205/Trp206) to an alanine residue, either individually or together, in silico revealed that the individual tryptophan residue's contribution to the tryptophan ‘anchor’ was additive. When the three tryptophan residues were mutated to alanine in vitro, the resulting mutant Top1 differed from wild-type Top1 in that it lacked processivity, exhibited resistance to camptothecin and was inactivated by urea. The results indicated that the tryptophan anchor stabilized the N-terminus of the functional domain and prevented the loss of Top1 structure and function.

scholarly journals The C-Terminal Flexible Domain of the Heme Chaperone CcmE Is Important but Not Essential for Its Function

2003 ◽  
Vol 185 (13) ◽  
pp. 3821-3827 ◽  
Author(s):  
Elisabeth Enggist ◽  
Linda Thöny-Meyer
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Heme Binding ◽  
Membrane Anchor ◽  
Terminal Domain ◽  
Conserved Domain ◽  
C Terminus ◽  
Helical Turn ◽  
Low Activity

ABSTRACT CcmE is a heme chaperone active in the cytochrome c maturation pathway of Escherichia coli. It first binds heme covalently to strictly conserved histidine H130 and subsequently delivers it to apo-cytochrome c. The recently solved structure of soluble CcmE revealed a compact core consisting of a β-barrel and a flexible C-terminal domain with a short α-helical turn. In order to elucidate the function of this poorly conserved domain, CcmE was truncated stepwise from the C terminus. Removal of all 29 amino acids up to crucial histidine 130 did not abolish heme binding completely. For detectable transfer of heme to type c cytochromes, only one additional residue, D131, was required, and for efficient cytochrome c maturation, the seven-residue sequence 131DENYTPP137 was required. When soluble forms of CcmE were expressed in the periplasm, the C-terminal domain had to be slightly longer to allow detection of holo-CcmE. Soluble full-length CcmE had low activity in cytochrome c maturation, indicating the importance of the N-terminal membrane anchor for the in vivo function of CcmE.

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