scholarly journals Structural characterization of the ribonuclease H-like type ASKHA superfamily kinase MK0840 fromMethanopyrus kandleri

2013 ◽  
Vol 69 (12) ◽  
pp. 2440-2450 ◽  
Author(s):  
Magdalena Schacherl ◽  
Sandro Waltersperger ◽  
Ulrich Baumann
Keyword(s):  
Conformational Changes ◽  
Metabolic Pathways ◽  
Bound States ◽  
Degradation Products ◽  
Nucleotide Binding ◽  
Active Site Residues ◽  
Methanopyrus Kandleri ◽  
Sugar Kinase ◽  
Actin Superfamily

Murein recycling is a process in which microorganisms recover peptidoglycan-degradation products in order to utilize them in cell wall biosynthesis or basic metabolic pathways. Methanogens such asMethanopyrus kandlericontain pseudomurein, which differs from bacterial murein in its composition and branching. Here, four crystal structures of the putative sugar kinase MK0840 fromM. kandleriin apo and nucleotide-bound states are reported. MK0840 shows high similarity to bacterial anhydro-N-acetylmuramic acid kinase, which is involved in murein recycling. The structure shares a common fold with panthothenate kinase and the 2-hydroxyglutaryl-CoA dehydratase component A, both of which are members of the ASKHA (acetate and sugar kinases/Hsc70/actin) superfamily of phosphotransferases. Local conformational changes in the nucleotide-binding site between the apo and holo forms are observed upon nucleotide binding. Further insight is given into domain movements and putative active-site residues are identified.

Structural and functional roles of dynamically correlated residues in thymidylate kinase

2018 ◽  
Vol 74 (4) ◽  
pp. 341-354 ◽  
Author(s):  
Santosh Kumar Chaudhary ◽  
Jeyaraman Jeyakanthan ◽  
Kanagaraj Sekar
Keyword(s):  
Molecular Dynamics ◽  
Conformational Changes ◽  
Active Site ◽  
Bound States ◽  
Catalytic Mechanism ◽  
Phosphoryl Transfer ◽  
Active Site Residues ◽  
Thymidylate Kinase ◽  
Binary And Ternary Complexes ◽  
Insight Into

Thymidylate kinase is an important enzyme in DNA synthesis. It catalyzes the conversion of thymidine monophosphate to thymidine diphosphate, with ATP as the preferred phosphoryl donor, in the presence of Mg2+. In this study, the dynamics of the active site and the communication paths between the substrates, ATP and TMP, are reported for thymidylate kinase fromThermus thermophilus. Conformational changes upon ligand binding and the path for communication between the substrates and the protein are important in understanding the catalytic mechanism of the enzyme. High-resolution X-ray crystal structures of thymidylate kinase in apo and ligand-bound states were solved. This is the first report of structures of binary and ternary complexes of thymidylate kinase with its natural substrates ATP and ATP–TMP, respectively. Distinct conformations of the active-site residues, the P-loop and the LID region observed in the apo and ligand-bound structures revealed that their concerted motion is required for the binding and proper positioning of the substrate TMP. Structural analyses provide an insight into the mode of substrate binding at the active site. The residues involved in communication between the substrates were identified through network analysis using molecular-dynamics simulations. The residues identified showed high sequence conservation across species. Biochemical analyses show that mutations of these residues either resulted in a loss of activity or affected the thermal stability of the protein. Further, molecular-dynamics analyses of mutants suggest that the proper positioning of TMP is important for catalysis. These data also provide an insight into the phosphoryl-transfer mechanism.

The Engines of Biomolecular Motors

2004 ◽  
Author(s):  
Jung-Chi Liao ◽  
George Oster
Keyword(s):  
Binding Site ◽  
Conformational Changes ◽  
Bound States ◽  
Circumferential Stress ◽  
Nucleotide Binding ◽  
Mechanical Work ◽  
Nucleoside Triphosphate ◽  
Biomolecular Motors ◽  
Adjacent Structures ◽  
Β Sheet

The majority of biomolecular motors are powered by nucleoside triphosphate (NTP), especially adenosine triphosphate (ATP). These motors consist of a β-sheet with highly conserved motifs and the nucleotide binding domain around it. The highly conserved protein folds are the engines of these motors, which convert the energy of NTP hydrolysis cycle to mechanical work. Although functions of molecular motors are widely diverse, (including cargo movement, DNA unwinding, protein degradation, ion pumping, etc), the nucleotide binding domains are very similar. In the binding site, NTP undergoes a hydrolysis cycle E+NTP⇄E·NTP⇄E•NTP⇄E•NDP•Pi⇄E•NDP+Pi⇄E+NDP+Pi where E is the enzyme (motor protein), the small dot represents the docking of NTP, and the large dot represents the tightly-bound states. The hydrogen bond network formed in the NTP binding step, as shown in Figure 1 [1], deforms the β-sheet and adjacent structures. The local deformation propagates to conformational changes of functional residues to do mechanical work or to change the affinity to the substrate [2]. For multimeric motor proteins, we must also consider the stress paths among subunits which control the sequence and the activity of the protein. Stress trajectories emanating from a binding site either passes through a circumferential stress loop or a stress loop through the substrate.

scholarly journals Atomic-level characterization of conformational transition and substrate binding of xCT transporter

2018 ◽  
Author(s):  
M. Sharma ◽  
A. C. Rohithaswa
Keyword(s):  
Ligand Binding ◽  
Conformational Changes ◽  
Bound States ◽  
Conformational Transition ◽  
Substrate Binding ◽  
Transmembrane Helices ◽  
Neurological Processes ◽  
Dynamics Simulations ◽  
Alternating Access

AbstractxCT is a component of heterodimeric amino acids transporter system Xc- that has been known to work at the cross-roads of maintaining neurological processes and regulating antioxidant defense. xCT is a sodium-independent amino acid antiporter, that imports L- cystine and exports L-glutamate in a 1:1 ratio. The transporter has 12 transmembrane domains with intracellular N- and C-termini, which can undergo various conformational changes while switching the ligand accessibilities from intracellular to extracellular site. In the present study, we generated two homology models of human xCT in two distinct conformations: inward facing occluded state and outward facing open state. We investigated the conformational transitions within these two states by employing series of targeted molecular dynamics simulations. Our results indicated the substrate translocation channel composed of transmembrane helices TMs 1, 3, 6, 8, and 10. Further, we analyzed the ligand binding within the intermediate conformations obtained from the transition simulations. We docked anionic L-cystine and L-glutamate within the cavities alone or in combination to assess the two distinct binding scenarios for xCT as antiporter. We also assessed the interactions between the ligand and xCT and observed that ligands bind to similar residues within the channel, and these residues are essential for substrate binding/permeation. In addition, we analyzed the correlations between ligand binding and conformational transition and observed conformations that are representatives for intermediate ligand bound states. The results presented in the study provide insights into the interplay of conformational transition and ligand binding as xCT goes from one probable conformation to another while transporting the ligand. And the data thus adds to the existing evidence of alternating access mechanism pertaining to the functioning of transporters.

Characterization of dissolved material during the initial phase of softwood kraft pulping

TAPPI Journal ◽  
2013 ◽  
Vol 12 (1) ◽  
pp. 37-43 ◽  
Author(s):  
HANNU PAKKANEN ◽  
TEEMU PALOHEIMO ◽  
RAIMO ALÉN
Keyword(s):  
Mass Transfer ◽  
Molecular Mass ◽  
Initial Phase ◽  
Degradation Products ◽  
Kraft Pulping ◽  
Reaction Products ◽  
Cooking Temperature ◽  
Molecular Masses ◽  
Aliphatic Carboxylic Acids

The influence of various cooking parameters, such as effective alkali, cooking temperature, and cooking time on the formation of high molecular mass lignin-derived and low molecular mass carbohydrates-derived (aliphatic carboxylic acids) degradation products, mainly during the initial phase of softwood kraft pulping was studied. In addition, the mass transfer of all of these degradation products was clarified based on their concentrations in the cooking liquor inside and outside of the chips. The results indicated that the degradation of the major hemicellulose component, galactoglucomannan, typically was dependent on temperature, and the maximum degradation amount was about 60%. In addition, about 60 min at 284°F (140°C) was needed for leveling off the concentrations of the characteristic reaction products (3,4-dideoxy-pentonic and glucoisosaccharinic acids) between these cooking liquors. Compared with low molecular mass aliphatic acids, the mass transfer of soluble lignin fragments with much higher molecular masses was clearly slower.

Synthesis and Characterization of Potential and Degraded Impurities of Regadenoson

2020 ◽  
Vol 16 (8) ◽  
pp. 1130-1139
Author(s):  
Singaram Sathiyanarayanan ◽  
Chidambaram Subramanian Venkatesan ◽  
Senthamaraikannan Kabilan
Keyword(s):  
Mass Spectra ◽  
Degradation Products ◽  
Hplc Method ◽  
Forced Degradation ◽  
Spectroscopic Techniques ◽  
A2a Adenosine Receptor ◽  
Degradation Study ◽  
Related Substances ◽  
Adenosine Receptor Agonist

Background: Regadenoson is an A2A adenosine receptor agonist that is a coronary vasodilator and commonly used as a pharmacologic cardiac stressing agents. Methods: HPLC method was used for the analysis of related substances. The degraded impurities during the process were isolated and characterized by IR, Mass and NMR spectral analysis. Results: Forced degradation study of regadenoson under conditions of hydrolysis (neutral, acidic and alkaline) and oxidations suggested in the ICH Q1A(R2) was accomplished. The drug showed significant degradation under all the above conditions. On the whole, five novel degradation products were found under diverse conditions along with process related impurities which were not reported earlier. Conclusion: All the degradation products were well characterized by using advanced spectroscopic techniques like IR, 1H NMR, 13C NMR and Mass spectra. The identification of these impurities will be productive for the quality control during the production and stability behavior of the regadenoson drug substance.

Characterization of Nucleotide Binding ­Site-Encoding Genes in Sweetpotato, Ipomoea batatas(L.) Lam., and Their Response to Biotic and Abiotic Stresses

2021 ◽  
pp. 1-15
Author(s):  
Zengzhi Si ◽  
Yake Qiao ◽  
Kai Zhang ◽  
Zhixin Ji ◽  
Jinling Han
Keyword(s):  
Binding Site ◽  
Abiotic Stresses ◽  
Ipomoea Batatas ◽  
Expression Profiles ◽  
Nucleotide Binding ◽  
Regulatory Elements ◽  
Nucleotide Binding Site ◽  
Biotic And Abiotic Stresses ◽  
Encoding Genes

Sweetpotato, <i>Ipomoea batatas</i> (L.) Lam., is an important and widely grown crop, yet its production is affected severely by biotic and abiotic stresses. The nucleotide binding site (NBS)-encoding genes have been shown to improve stress tolerance in several plant species. However, the characterization of NBS-encoding genes in sweetpotato is not well-documented to date. In this study, a comprehensive analysis of NBS-encoding genes has been conducted on this species by using bioinformatics and molecular biology methods. A total of 315 NBS-encoding genes were identified, and 260 of them contained all essential conserved domains while 55 genes were truncated. Based on domain architectures, the 260 NBS-encoding genes were grouped into 6 distinct categories. Phylogenetic analysis grouped these genes into 3 classes: TIR, CC (I), and CC (II). Chromosome location analysis revealed that the distribution of NBS-encoding genes in chromosomes was uneven, with a number ranging from 1 to 34. Multiple stress-related regulatory elements were detected in the promoters, and the NBS-encoding genes’ expression profiles under biotic and abiotic stresses were obtained. According to the bioinformatics analysis, 9 genes were selected for RT-qPCR analysis. The results revealed that <i>IbNBS75</i>, <i>IbNBS219</i>, and <i>IbNBS256</i> respond to stem nematode infection; <i>Ib­NBS240</i>, <i>IbNBS90</i>, and <i>IbNBS80</i> respond to cold stress, while <i>IbNBS208</i>, <i>IbNBS71</i>, and <i>IbNBS159</i> respond to 30% PEG treatment. We hope these results will provide new insights into the evolution of NBS-encoding genes in the sweetpotato genome and contribute to the molecular breeding of sweetpotato in the future.

scholarly journals Structural basis for activation and allosteric modulation of full-length calcium-sensing receptor

2021 ◽  
Vol 7 (23) ◽  
pp. eabg1483
Author(s):  
Tianlei Wen ◽  
Ziyu Wang ◽  
Xiaozhe Chen ◽  
Yue Ren ◽  
Xuhang Lu ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Bound States ◽  
Hormone Secretion ◽  
Allosteric Modulation ◽  
Full Length ◽  
Structural Basis ◽  
Endocrine Disorders ◽  
Calcium Sensing Receptor ◽  
Calcium Sensing ◽  
Class C

Calcium-sensing receptor (CaSR) is a class C G protein–coupled receptor (GPCR) that plays an important role in calcium homeostasis and parathyroid hormone secretion. Here, we present multiple cryo–electron microscopy structures of full-length CaSR in distinct ligand-bound states. Ligands (Ca2+ and l-tryptophan) bind to the extracellular domain of CaSR and induce large-scale conformational changes, leading to the closure of two heptahelical transmembrane domains (7TMDs) for activation. The positive modulator (evocalcet) and the negative allosteric modulator (NPS-2143) occupy the similar binding pocket in 7TMD. The binding of NPS-2143 causes a considerable rearrangement of two 7TMDs, forming an inactivated TM6/TM6 interface. Moreover, a total of 305 disease-causing missense mutations of CaSR have been mapped to the structure in the active state, creating hotspot maps of five clinical endocrine disorders. Our results provide a structural framework for understanding the activation, allosteric modulation mechanism, and disease therapy for class C GPCRs.

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