An Efficient System for 30-K MAS-NMR-DNP for Macromolecule Structure Determinations

Doty Scientific, Inc., Columbia SC

Investigators

Abstract

The critical importance of solid-state NMR (ssNMR) was recently demonstrated by, after nearly two decades of intense efforts, yielding the first atomic-resolution structures of the A?40 and A?42 amyloid fibrils that play a crucial role in Alzheimer?s Disease (AD). Key to that advance is a technique denoted as dynamic nuclear polarization (DNP) with magic angle spinning (MAS). For technical reasons (discussed below) these experiments have been carried out temperatures on the order of 100 K. While Cryo EM, scanning tunneling electron microscopy (STEM) and other methods provided useful information, recent advances in MAS-NMR methods have refined the structures to atomic resolution by adding 487 NMR constraints, including sidechain dynamics important in protein functions and in understanding of myriad mechanisms of their action. Moreover, only a small number of insoluble membrane proteins larger than 35 kDa with backbone and side-chain assignments completed by >80% have been deposited into the Protein Data Bank. Hence, developing transformational advances for ssNMR is crucial for both structural biology and biomedical research in general, for progress in curing Alzheimer?s Disease and cancer, and for providing regio-specific drug binding information enabling detailing of the mechanism of action for effective drugs. This proposed Phase I SBIR will show feasibility of revolutionary advances in ultra-compact super-effective cryogenic heat exchangers that are small enough to fit inside MAS-DNP probes that, with the novel cryogenic system developments planned for Phase-II, will enable routine low-cost MAS-DNP down to 30 K. This will dramatically reduce the amount of microwave power required for the electron spin saturation needed for the DNP method, and will thereby enable MAS-DNP using advanced low-cost THz solid-state sources rather than expensive gyrotrons. The Phase-II MAS-DNP system will have zero consumption of liquid cryogens, negligible helium gas consumption (only during sample changes), affordable equipment costs, and operating costs of only a few dollars per hour.