scholarly journals Assignment of the resting state of the secondary active transporter BetP by integrating spectroscopic measurements and molecular simulations

2018 ◽  
Author(s):  
Vanessa Leone ◽  
Izabela Waclawska ◽  
Caroline Koshy ◽  
Katharina Kossman ◽  
Monika Sharma ◽  
...  
Keyword(s):  
Double Resonance ◽  
Molecular Simulations ◽  
Enhanced Sampling ◽  
Experimental Conditions ◽  
Physiological Conditions ◽  
Data Comparison ◽  
Osmotic Stress Response ◽  
Biochemical Studies ◽  
Transport Cycle ◽  
Crystallographic Structures

The glycine betaine symporter BetP regulates the osmotic stress response of Corynebacterium glutamicum, a soil bacterium used extensively in biotechnology. Although BetP is a homotrimer, biochemical studies have shown that each protomer is able to transport its substrate independently. Crystallographic structures of BetP have been determined in several conformations, seemingly capturing outward-open, inward-open and occluded states, both loaded with the substrate and in the apo form. However, it has been challenging to establish a correspondence between each of these structures and specific states in the mechanism of the transporter under more physiological conditions. To this end, we examined the dynamics of spin-labelled BetP using pulsed electron-electron double resonance (PELDOR) under different stimuli. We then carried out molecular simulations of structures of the BetP monomer to interpret the PELDOR data, using the enhanced-sampling methodology EBMetaD [Marinelli & Faraldo-Gomez, 2015, Biophys J 108(12):2779-2782], whereby the dynamics of the protein are minimally biased so as to reproduce the experimental data. Comparison of the magnitude of the biasing work required for different input structures permitted us to assign them to specific states of the transport cycle under each of the experimental conditions. In particular, this analysis showed that BetP adopts inward-facing conformations in the presence of excess sodium, and a mixture of states when betaine is added. These studies better delineate the major conformations adopted by BetP in its transport cycle, and therefore provide important insights into its mechanism. More broadly, we illustrate how integrative simulations can aid interpretation of ambiguous structural and spectroscopic data on membrane proteins.

scholarly journals Extended Phase-Space Methods for Enhanced Sampling in Molecular Simulations: A Review

2015 ◽  
Vol 3 ◽  
Author(s):  
Hiroshi Fujisaki ◽  
Kei Moritsugu ◽  
Yasuhiro Matsunaga ◽  
Tetsuya Morishita ◽  
Luca Maragliano
Keyword(s):  
Phase Space ◽  
Molecular Simulations ◽  
Extended Phase Space ◽  
Enhanced Sampling ◽  
Extended Phase ◽  
Phase Space Methods

scholarly journals Conformational ensemble of human α-synuclein physiological form predicted by molecular simulations

2016 ◽  
Vol 18 (8) ◽  
pp. 5702-5706 ◽  
Author(s):  
G. Rossetti ◽  
F. Musiani ◽  
E. Abad ◽  
D. Dibenedetto ◽  
H. Mouhib ◽  
...  
Keyword(s):  
Protein Dynamics ◽  
Molecular Simulations ◽  
Intramolecular Interactions ◽  
Conformational Ensemble ◽  
Enhanced Sampling ◽  
Post Translational Modification ◽  
Α Helix

Enhanced sampling simulations of N-terminally acetylated human α-synuclein suggest that the post-translational modification leads to the formation of a transient amphipathic α-helix altering protein dynamics at the N-terminal and intramolecular interactions.

Lanthanum in Human Red Cell

1982 ◽  
Vol 40 ◽  
pp. 280-281
Author(s):  
Ezzatollah Keyhani ◽  
Eva Bland
Keyword(s):  
Sialic Acid ◽  
Ultrastructural Localization ◽  
Cellular Membrane ◽  
Red Cells ◽  
Red Cell ◽  
Total Lipids ◽  
Experimental Conditions ◽  
Sialic Acid Content ◽  
Biochemical Studies ◽  
Human Red Cells

The human red cell contains four major glycoproteins : sialoglycoprotein or PAS 1; Band III; PAS 2 and PAS 3. With the exception of Band III, all other glycoproteins contain high levels of sialic acid. In addition, human red cells contain glycolipids which have been estimated to be less than 5% of the total lipids. Lanthanum has been used as tracer of extracellular space and as a probe for studies of Ca++ metabolism across various types of cellular membrane. Biochemical studies led to the conclusion that lanthanum inhibition of Ca++ metabolism was related to the sialic acid content of the cell membrane. The purpose of this communication is to describe the ultrastructural localization of lanthanum in human red cells under various experimental conditions.

scholarly journals Biomolecular conformational changes and ligand binding: from kinetics to thermodynamics

2017 ◽  
Vol 8 (9) ◽  
pp. 6466-6473 ◽  
Author(s):  
Yong Wang ◽  
João Miguel Martins ◽  
Kresten Lindorff-Larsen
Keyword(s):  
Ligand Binding ◽  
Conformational Changes ◽  
Molecular Simulations ◽  
Kinetic Properties ◽  
Enhanced Sampling ◽  
Biomolecular Systems

Biomolecular systems such as protein–ligand complexes are governed by thermodynamic and kinetic properties that may be estimated at the same time through enhanced-sampling molecular simulations.

Monitoring of blood gas parameters and acid-base balance of pregnant and non-pregnant rabbits (Oryctolagus cuniculus) in routine experimental conditions

1992 ◽  
Vol 26 (2) ◽  
pp. 73-79 ◽  
Author(s):  
M. M. Barzago ◽  
A. Bortolotti ◽  
D. Omarini ◽  
J. J. Aramayona ◽  
M. Bonati
Keyword(s):  
New Zealand ◽  
Oryctolagus Cuniculus ◽  
Housing Conditions ◽  
Blood Gas ◽  
Acid Base Balance ◽  
Acid Base ◽  
Experimental Conditions ◽  
Physiological Conditions ◽  
Base Balance ◽  
Blood Gas Parameters

Blood gas parameters and acid-base balance values were determined in adult pregnant New Zealand rabbits ( Oryctolagus cuniculus) in standard laboratory housing conditions and during anaesthesia with an association of ketamine-chlorpromazine, administered before surgical procedures. All the variables were also studied in adult non-pregnant female, used as controls. No differences in pH, sO2c, O2Hb, COHb, sO2m and a-vDO2 were found between pregnant and non-pregnant rabbits in physiological conditions and during anaesthesia. Ketamine-chlorpromazine and pregnancy seemed to change the other parameters used to assess the acid-base balance and the oxygenation conditions. Anaesthesia affected only Hb, O2Ct, O2Cap, CcO2 and P50. The additive effect of pregnancy and anaesthesia modified pCO2, pO2, HCO3-, TCO2, BEb, SBC, BEecf, A-aDO2, RI, MetHb, RHb, CaO2 and CvO2. The patterns described are close to those of other species, suggesting the New Zealand rabbit might be a reliable animal model for monitoring selected variables.

scholarly journals Meta-analysis of gene expression profiles in granulosa cells during folliculogenesis

Reproduction ◽  
2016 ◽  
Vol 151 (6) ◽  
pp. R103-R110 ◽  
Author(s):  
Daulat Raheem Khan ◽  
Éric Fournier ◽  
Isabelle Dufort ◽  
François J Richard ◽  
Jaswant Singh ◽  
...  
Keyword(s):  
Gene Expression ◽  
Granulosa Cell ◽  
Granulosa Cells ◽  
Expression Profiles ◽  
Post Partum ◽  
Meta Analysis ◽  
Expression Patterns ◽  
Comprehensive Overview ◽  
Experimental Conditions ◽  
Physiological Conditions

Abstract Folliculogenesis involves coordinated profound changes in different follicular compartments and significant modifications of their gene expression patterns, particularly in granulosa cells. Huge datasets have accumulated from the analyses of granulosa cell transcriptomic signatures in predefined physiological contexts using different technological platforms. However, no comprehensive overview of folliculogenesis is available. This would require integration of datasets from numerous individual studies. A prerequisite for such integration would be the use of comparable platforms and experimental conditions. The EmbryoGENE program was created to study bovine granulosa cell transcriptomics under different physiological conditions using the same platform. Based on the data thus generated so far, we present here an interactive web interface called GranulosaIMAGE (Integrative Meta-Analysis of Gene Expression), which provides dynamic expression profiles of any gene of interest and all isoforms thereof in granulosa cells at different stages of folliculogenesis. GranulosaIMAGE features two kinds of expression profiles: gene expression kinetics during bovine folliculogenesis from small (6 mm) to pre-ovulatory follicles under different hormonal and physiological conditions and expression profiles of granulosa cells of dominant follicles from post-partum cows in different metabolic states. This article provides selected examples of expression patterns along with suggestions for users to access and generate their own patterns using GranulosaIMAGE. The possibility of analysing gene expression dynamics during the late stages of folliculogenesis in a mono-ovulatory species such as bovine should provide a new and enriched perspective on ovarian physiology.

scholarly journals Conformational flexibility in the catalytic triad revealed by the high-resolution crystal structure ofStreptomyces erythraeustrypsin in an unliganded state

2014 ◽  
Vol 70 (3) ◽  
pp. 833-840 ◽  
Author(s):  
Elise Blankenship ◽  
Krishna Vukoti ◽  
Masaru Miyagi ◽  
David T. Lodowski
Keyword(s):  
Serine Proteases ◽  
Catalytic Mechanism ◽  
Data Bank ◽  
Catalytic Triad ◽  
Detailed Structural Analysis ◽  
Structural Rigidity ◽  
Biochemical Studies ◽  
Crystallographic Structures ◽  
Extreme Stability ◽  
Resolution Structure

With more than 500 crystal structures determined, serine proteases make up greater than one-third of all proteases structurally examined to date, making them among the best biochemically and structurally characterized enzymes. Despite the numerous crystallographic and biochemical studies of trypsin and related serine proteases, there are still considerable shortcomings in the understanding of their catalytic mechanism.Streptomyces erythraeustrypsin (SET) does not exhibit autolysis and crystallizes readily at physiological pH; hence, it is well suited for structural studies aimed at extending the understanding of the catalytic mechanism of serine proteases. While X-ray crystallographic structures of this enzyme have been reported, no coordinates have ever been made available in the Protein Data Bank. Based on this, and observations on the extreme stability and unique properties of this particular trypsin, it was decided to crystallize it and determine its structure. Here, the first sub-angstrom resolution structure of an unmodified, unliganded trypsin crystallized at physiological pH is reported. Detailed structural analysis reveals the geometry and structural rigidity of the catalytic triad in the unoccupied active site and comparison to related serine proteases provides a context for interpretation of biochemical studies of catalytic mechanism and activity.

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