Session TOB. There are 5 abstracts in this session.



Session: BIOMOLECULAR 2 - NMR IN SOLIDS, time: 08:30 - 8:55 am

Oligomeric Proteins in Cell Signaling 


Ann Mcdermott
Columbia University, New York, NY

Many oligomeric proteins have been implicated in cell signaling in human health and disease. The oligomeric RIPK1:RIPK3 complex (necrosome) signals necroptosis in the context of immune defense, cancer and neurodegenerative diseases. Using solid-state NMR, we determined the high-resolution structure of the necrosome core. RIPK1 and RIPK3 assume a new serpentine hetero-amyloid fold with stabilizing elements analogous to other amyloids including a hydrophobic core with both hetero and homo hydrophobic contacts, and solvent exposed “ladders” of interacting amino acids. The talk will also contrast this system to other oligomeric signaling complexes.


Session: BIOMOLECULAR 2 - NMR IN SOLIDS, time: 08:55 - 9:20 am

Leveraging Solid-state NMR Techniques to Characterize Host Defense Metallopeptides Active at Lipid Bilayers


Alexander Greenwood1; Robert Vold1; Ella Mihailescu2; Sergei Sukharev3; Mary Rooney1; Yawei Xiong1; Steven Paredes1; Jessica Hill1; Riqiang Fu4; Craig Bayse5; Myriam Cotten1
1William and Mary, Williamsburg, Virginia ; 2University of Maryland, Rockville, MD; 3University of Maryland, College Park, MD; 4National High Magnetic Field Laboratory, Tallahassee, FL; 5Old Dominion University, Norfolk, VA

Pathogenic membranes are battlegrounds for antimicrobial compounds, including host defense peptides (HDPs) and Cu2+. Conventionally, HDPs are known for physically disrupting membranes and/or crossing them to access intracellular targets. Our group explores the new paradigm that piscidins, which are histidine-rich HDPs, sensitize membranes for physical disruption by exploiting membrane heterogeneity, and using Cu2+-coordination to modulate peptide charge and conformation, and chemically damage lipids. We present data linking several functional and physicochemical characteristics of the piscidins. We show that their membranolytic effects, pH-behaviors, modulation of mechanosensitive-channel activation, and effects on bacterial cell division can be correlated to their modulation of charge and conformation as a function of histidine content and metallation state, directionality of membrane insertion, and exploitation of membrane heterogeneity.


Session: BIOMOLECULAR 2 - NMR IN SOLIDS, time: 09:20 - 9:35 am

Simultaneous and parallel acquisition for protein resonance assignment in solid-state NMR


Angelo Gallo; W. Trent Franks; Józef R. Lewandowski
University of Warwick, Coventry, United Kingdom

In this contribution, we present applications of dual-receiver parallel acquisition suite of experiments to obtain multiple 3D spectra involving different combinations of 1H, 13C and 15N correlations in a single experiment. The unique feature of the experiment presented here, compared to the previously reported dual or multiple receiver experiments, is the introduction of different chemical shifts during the second indirect dimension evolution.


Session: BIOMOLECULAR 2 - NMR IN SOLIDS, time: 09:35 - 9:50 am

Automated backbone resonance assignment of large proteins using redundant linking from a single simultaneous acquisition


Jan Stanek1; Tobias Schubeis2; Piotr Paluch1
1University of Warsaw, Warsaw, Poland; 2FRE 2034 (CNRS/UCB Lyon1/ENS Lyon), Villeurbanne, France

Resonance assignment in magic-angle spinning (MAS) NMR remains a bottleneck in protein structural studies, and even the state-of-the-art protocols fail for proteins above ~20 kDa. Here we leverage the benefits of 1H detection, fast (110 kHz) MAS and high magnetic fields to design an approach that lifts this limitation. Through the creation, coevolution and acquisition of independent magnetization pathways, a single self-consistent dataset can be acquired, providing a leap forward in sensitivity, and highly redundant linking that supports fully-automated peak picking and sequential resonance assignment. The method is demonstrated with the assignment of the largest protein to date in the solid state, the 42.5 kDa maltose binding protein, using a single fully-protonated  sample and a week of spectrometer time.


Session: BIOMOLECULAR 2 - NMR IN SOLIDS, time: 09:50 - 10:05 am

Characterization of Phosphatidylinositol Phosphates Binding to Pleckstrin Homology Domains in Lipid Bilayers by Solid-state NMR


Andrew Nieuwkoop; Jacqueline Perodeau; Ashley Bernstein; Robert Palmere; Stefany Lazieh
Rutgers University, Piscataway, NJ

Phosphatidylinositol phosphates (PIPs) are a class of membrane lipid that regulate diverse cell processes in eukaryotic organisms. The presence of specific PIP species helps determine the spatial and temporal location, as well as the activity, of hundreds of proteins containing diverse PIP binding domains. This work aims to use solid-state NMR to understand the properties of PIP containing lipid bilayers, map the PIP binding sites of pleckstrin homology (PH) domains, a class of PIP binders, and characterize the effects of PIP binding on the structure and dynamics of PH domains. We utilize 13C, 15N, 31P and 1H detected solid-state NMR and MAS rates ranging from 10 to 100+ kHz.