Isotopic Effects on Intermolecular and Intramolecular Structure and Dynamics in Hydrogen, Deuterium, and Tritium Liquids: Normal Liquid and Weakly and Strongly Cooled Liquids

2018 ◽  
Vol 122 (34) ◽  
pp. 8233-8242 ◽  
Author(s):  
Kiharu Abe ◽  
Shutaro Yamaoka ◽  
Kim Hyeon-Deuk
Keyword(s):  
Normal Liquid ◽  
Structure And Dynamics ◽  
Hydrogen Deuterium ◽  
Isotopic Effects

scholarly journals Insights into HIV-1 proviral transcription from integrative structure and dynamics of the Tat:AFF4:P-TEFb:TAR complex

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Ursula Schulze-Gahmen ◽  
Ignacia Echeverria ◽  
Goran Stjepanovic ◽  
Yun Bai ◽  
Huasong Lu ◽  
...  
Keyword(s):  
Major Groove ◽  
Binding Model ◽  
Saxs Data ◽  
Recognition Motif ◽  
Structure And Dynamics ◽  
Hydrogen Deuterium ◽  
Central Loop ◽  
Binding Loop ◽  
Hiv 1 ◽  
Arginine Rich Motif

HIV-1 Tat hijacks the human superelongation complex (SEC) to promote proviral transcription. Here we report the 5.9 Å structure of HIV-1 TAR in complex with HIV-1 Tat and human AFF4, CDK9, and CycT1. The TAR central loop contacts the CycT1 Tat-TAR recognition motif (TRM) and the second Tat Zn2+-binding loop. Hydrogen-deuterium exchange (HDX) shows that AFF4 helix 2 is stabilized in the TAR complex despite not touching the RNA, explaining how it enhances TAR binding to the SEC 50-fold. RNA SHAPE and SAXS data were used to help model the extended (Tat Arginine-Rich Motif) ARM, which enters the TAR major groove between the bulge and the central loop. The structure and functional assays collectively support an integrative structure and a bipartite binding model, wherein the TAR central loop engages the CycT1 TRM and compact core of Tat, while the TAR major groove interacts with the extended Tat ARM.

scholarly journals Structure and dynamics of an α-fucosidase reveal a mechanism for highly efficient IgG transfucosylation

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Erik H. Klontz ◽  
Chao Li ◽  
Kyle Kihn ◽  
James K. Fields ◽  
Dorothy Beckett ◽  
...  
Keyword(s):  
Active Site ◽  
Lactobacillus Casei ◽  
Molecular Mechanisms ◽  
Dynamic Simulations ◽  
Effector Functions ◽  
Exchange Mass Spectrometry ◽  
Structure And Dynamics ◽  
Hydrogen Deuterium ◽  
Deuterium Exchange Mass Spectrometry ◽  
Core Fucosylation

AbstractFucosylation is important for the function of many proteins with biotechnical and medical applications. Alpha-fucosidases comprise a large enzyme family that recognizes fucosylated substrates with diverse α-linkages on these proteins. Lactobacillus casei produces an α-fucosidase, called AlfC, with specificity towards α(1,6)-fucose, the only linkage found in human N-glycan core fucosylation. AlfC and certain point mutants thereof have been used to add and remove fucose from monoclonal antibody N-glycans, with significant impacts on their effector functions. Despite the potential uses for AlfC, little is known about its mechanism. Here, we present crystal structures of AlfC, combined with mutational and kinetic analyses, hydrogen–deuterium exchange mass spectrometry, molecular dynamic simulations, and transfucosylation experiments to define the molecular mechanisms of the activities of AlfC and its transfucosidase mutants. Our results indicate that AlfC creates an aromatic subsite adjacent to the active site that specifically accommodates GlcNAc in α(1,6)-linkages, suggest that enzymatic activity is controlled by distinct open and closed conformations of an active-site loop, with certain mutations shifting the equilibrium towards open conformations to promote transfucosylation over hydrolysis, and provide a potentially generalizable framework for the rational creation of AlfC transfucosidase mutants.

scholarly journals Structure and Dynamics of a Site-Specific Labeled Fc Fragment with Altered Effector Functions

Pharmaceutics ◽  
2019 ◽  
Vol 11 (10) ◽  
pp. 546 ◽  
Author(s):  
D. Travis Gallagher ◽  
Chris McCullough ◽  
Robert G. Brinson ◽  
Joomi Ahn ◽  
John P. Marino ◽  
...  
Keyword(s):  
Mass Spectroscopy ◽  
Insertion Site ◽  
Deuterium Exchange ◽  
Energy Estimates ◽  
Hydrogen Deuterium Exchange ◽  
Site Specific ◽  
Structure And Dynamics ◽  
Drug Conjugates ◽  
Backward Shift ◽  
Hydrogen Deuterium

Antibody-drug conjugates (ADCs) are a class of biotherapeutic drugs designed as targeted therapies for the treatment of cancer. Among the challenges in generating an effective ADC is the choice of an effective conjugation site on the IgG. One common method to prepare site-specific ADCs is to engineer solvent-accessible cysteine residues into antibodies. Here, we used X-ray diffraction and hydrogen-deuterium exchange mass spectroscopy to analyze the structure and dynamics of such a construct where a cysteine has been inserted after Ser 239 (Fc-239i) in the antibody heavy chain sequence. The crystal structure of this Fc-C239i variant at 0.23 nm resolution shows that the inserted cysteine structurally replaces Ser 239 and that this causes a domino-like backward shift of the local polypeptide, pushing Pro 238 out into the hinge. Proline is unable to substitute conformationally for the wild-type glycine at this position, providing a structural reason for the previously observed abolition of both FcγR binding and antibody-dependent cellular cytotoxicity. Energy estimates for the both the FcγR interface (7 kcal/mol) and for the differential conformation of proline (20 kcal/mol) are consistent with the observed disruption of FcγR binding, providing a quantifiable case where strain at a single residue appears to disrupt a key biological function. Conversely, the structure of Fc-C239i is relatively unchanged at the intersection of the CH2 and CH3 domains; the site known to be involved in binding of the neonatal Fc receptor (FcRn), and an alignment of the Fc-C239i structure with an Fc structure in a ternary Fc:FcRn:HSA (human serum albumin) complex implies that these favorable contacts would be maintained. Hydrogen deuterium exchange mass spectroscopy (HDX-MS) data further suggest a significant increase in conformational mobility for the Fc-C239i protein relative to Fc that is evident even far from the insertion site but still largely confined to the CH2 domain. Together, the findings provide a detailed structural and dynamic basis for previously observed changes in ADC functional binding to FcγR, which may guide further development of ADC designs.

scholarly journals Interplay of disordered and ordered regions of a human small heat shock protein yields an ensemble of ‘quasi-ordered’ states

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Amanda F Clouser ◽  
Hannah ER Baughman ◽  
Benjamin Basanta ◽  
Miklos Guttman ◽  
Abhinav Nath ◽  
...  
Keyword(s):  
Heat Shock ◽  
Hybrid Approach ◽  
Small Heat Shock Protein ◽  
Exchange Mass Spectrometry ◽  
Structure And Dynamics ◽  
Hydrogen Deuterium ◽  
Deuterium Exchange Mass Spectrometry ◽  
Shock Proteins ◽  
Structural Insights

Small heat shock proteins (sHSPs) are nature’s ‘first responders’ to cellular stress, interacting with affected proteins to prevent their aggregation. Little is known about sHSP structure beyond its structured α-crystallin domain (ACD), which is flanked by disordered regions. In the human sHSP HSPB1, the disordered N-terminal region (NTR) represents nearly 50% of the sequence. Here, we present a hybrid approach involving NMR, hydrogen-deuterium exchange mass spectrometry, and modeling to provide the first residue-level characterization of the NTR. The results support a model in which multiple grooves on the ACD interact with specific NTR regions, creating an ensemble of ‘quasi-ordered’ NTR states that can give rise to the known heterogeneity and plasticity of HSPB1. Phosphorylation-dependent interactions inform a mechanism by which HSPB1 is activated under stress conditions. Additionally, we examine the effects of disease-associated NTR mutations on HSPB1 structure and dynamics, leveraging our emerging structural insights.

scholarly journals Interplay of disordered and ordered regions of a human small heat shock protein yields an ensemble of “quasi-ordered” states

2019 ◽  
Author(s):  
Amanda F. Clouser ◽  
Hannah E.R. Baughman ◽  
Benjamin Basanta ◽  
Miklos Guttman ◽  
Abhinav Nath ◽  
...  
Keyword(s):  
Heat Shock ◽  
Hybrid Approach ◽  
Small Heat Shock Protein ◽  
Exchange Mass Spectrometry ◽  
Structure And Dynamics ◽  
Hydrogen Deuterium ◽  
Deuterium Exchange Mass Spectrometry ◽  
Shock Proteins ◽  
Structural Insights

ABSTRACTSmall heat shock proteins (sHPSs) are nature’s “first responders” to cellular stress, interacting with affected proteins to prevent their aggregation. Little is known about sHSP structure beyond its structured α-crystallin domain (ACD), which is flanked by disordered regions. In the human sHSP HSPB1, the disordered N-terminal region (NTR) represents nearly 50% of the sequence. Here, we present a hybrid approach involving NMR, hydrogen-deuterium exchange mass spectrometry, and modeling to provide the first residue-level characterization of the NTR. The results support a model in which multiple grooves on the ACD interact with specific NTR regions, creating an ensemble of “quasi-ordered” NTR states that can give rise to the known heterogeneity and plasticity of HSPB1. Phosphorylation-dependent interactions inform a mechanism by which HSPB1 is activated under stress conditions. Additionally, we examine the effects of disease-associated NTR mutations on HSPB1 structure and dynamics, leveraging our emerging structural insights.

Nucleosome dynamics

2006 ◽  
Vol 73 ◽  
pp. 109-119 ◽  
Author(s):  
Chris Stockdale ◽  
Michael Bruno ◽  
Helder Ferreira ◽  
Elisa Garcia-Wilson ◽  
Nicola Wiechens ◽  
...  
Keyword(s):  
High Resolution ◽  
Chromatin Structure ◽  
Current Knowledge ◽  
Dynamic Properties ◽  
Structure And Dynamics ◽  
Nucleosome Dynamics ◽  
Sharp Focus ◽  
Recent Advances ◽  
Insight Into ◽  
Chromatin Structure And Dynamics

In the 30 years since the discovery of the nucleosome, our picture of it has come into sharp focus. The recent high-resolution structures have provided a wealth of insight into the function of the nucleosome, but they are inherently static. Our current knowledge of how nucleosomes can be reconfigured dynamically is at a much earlier stage. Here, recent advances in the understanding of chromatin structure and dynamics are highlighted. The ways in which different modes of nucleosome reconfiguration are likely to influence each other are discussed, and some of the factors likely to regulate the dynamic properties of nucleosomes are considered.

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