Dynamic Nuclear Polarization enhanced MAS ssNMR (DNP Workshop)

Next offering: In-person meetings were cancelled throughout 2020-2022 due to the pandemic; next possible offering is February 2024 (3-4 days)

Location: NHMFL Main Facility, Tallahassee, FL
Leaders: Joanna Long & Frederic Mentink-Vigier (TDP2)
Last Offered: October 22-24, 2019 – Find a summary here

In its initial offering (October 2019), this workshop focused on current biomolecular solid-state NMR strategies and making samples for DNP using conventional biradical additives for sensitivity enhancement. As this workshop matures, an added component focusing on targeted spin labeling along with rapid freezing approaches will be developed through guidance provided by our DBP partners Len Mueller, Tatyana Polenova, and Yoshitaki Ishii.  Participants will also be introduced to various software packages developed for structural biology applications with an emphasis on using ssNMR data as well as databases for depositing NMR data and best practices for disseminating data and data sharing.

Goals:

• Learn best practices for biomolecular applications of solid state NMR
• Develop a working knowledge of DNP hardware and strategies for radical introduction
• Appreciate the various factors affecting sensitivity and resolution at high magnetic fields for conventional and DNP enhanced biomolecular MAS ssNMR applications

Sample schedule:

Before you arrive: Registration/setup and safety training

Day 1: Introduction to biomolecular MAS ssNMR

• Tour of NHMFL and DNP Lab; History of DNP
• Lectures on basics of spin interactions and ssNMR, magic angle spinning, MAS ssNMR for protein struc­ture determination, isotopic enrichment strate­gies, ba­sics of an NMR probe circuit
• Focus on the probe, practicum on sample preparation, sample packing and magic angle spinning, ba­sics of probe tuning and matching, ¹³C CPMAS acquisition, evaluation of sensi­tiv­ity and resolution, calibration of pulse lengths and power levels

Day 2: Applications of biomolecular ssNMR for structure determination

• DNP mechanisms, an introduction to EPR for DNP practition­ers, approaches for introduction of stable, local­ized electron spins, DNP hardware, strategies for recapturing resolution at low temperatures
• Practicum on setting up multidimensional ssNMR experiments, analyzing ssNMR data for a model crystal­line protein

Day 3: Dynamic nuclear polarization

• Lectures on multidimensional NMR, ssNMR experiments for sequential assign­ments of proteins, ssNMR experiments for structural constraints, fac­tors affect­ing resolution
• Practicum on DNP sample preparation, low field EPR measure­ments, optimization of DNP polarization, meas­uring enhance­ments, evaluating resolution
• Sample analyses and discussion of results