The objective of this proposal is to carry out the scientific testing associated with a new NMR software package developed currently at Biosym, namely an integrated theoretical/computational tool for determining molecular structures from NMR data. The scheme combines an elaborate NMR Relaxation Matrix Approach (IRMA) with various molecular modeling formulations such as Distance Geometry (DG), Restrained Molecular Dynamics (RMD)/Restrained Energy Minimization (REM) and Dynamical Simulated Annealing (DSA). Structure determination proceeds by first `generating' a structure, with an extended version of T. Havel's metric matrix formulation called DG II or with a recently proposed DSA scheme, which has been implemented at Biosym using the molecular dynamics (MD) package DISCOVER. This is followed by structure refinement, pursued using a DISCOVER-integrated version of IRMA. The integrity and validity of this complex system is contingent upon careful scientific testing aimed at assessing limitations (on one hand) and relative advantages of alternative components (on the other). Moreover, the unique scientific rigor inherent in this system will be fully appreciated only if we can illustrate its ability to solve momentarily ambiguous molecular structures. DG II and the DISCOVER-based DSA will be evaluated comparatively using as test cases crambin and ZnIV fingerlike domain. Limitations and benefits of the two techniques will be pinpointed by subjecting bovine pancreatic trypsin inhibitor (BPTI) to a detailed investigation and comparing the results with previous studies. A sensitivity test of IRMA will be performed using the alpha-amylase inhibitor Tendamistat. The important implications of treating internal motions, to the extent that previous structural ambiguities can be resolved by properly accounted for dynamic phenomena, will be illustrated by re-investigating Neutrophil Peptide 5. The merit of this research in terms of technological innovation and commercial value will be borne out by a scientifically sound, novel, unique and very useful tool for elucidating NMR structures.
