The anti-human leukocyte antigen (HLA) class I monoclonal antibody (mAb) TP25.99 has a unique specificity since it recognizes both a conformational and a linear determinant expressed on the β2-μ-associated and β2-μ-free HLA class I heavy chains, respectively. Previously, we reported the identification of a cyclic and a linear peptide that inhibits mAb TP25.99 binding to the β2-μ-associated and β2-μ-free HLA class I heavy chains (S. A. Desai, X. Wang, E. J. Noronha, Q. Zhou, V. Rebmann, H. Grosse-Wilde, F. J. Moy, R. Powers, and S. Ferrone, submitted for publication). The linear X19 and cyclic LX-8 peptides contain sequence homologous to residues 239–242, 245, and 246 and to residues 194–198, respectively, of HLA class I heavy chain α3 domain. Analysis by two-dimensional transfer nuclear Overhauser effect spectroscopy of the induced solution structures of the linear X19 and cyclic LX-8 peptides in the presence of mAb TP25.99 showed that the two peptides adopt a similar structural motif despite the lack of sequence homology. The backbone fold is suggestive of a short helical segment followed by a tight turn, reminiscent of the determinant loop region (residues 194–198) on β2-μ-associated HLA class I heavy chains. The structural similarity between the linear X19 and cyclic LX-8 peptides and the lack of sequence homology suggests that mAb TP25.99 predominantly recognizes a structural motif instead of a consensus sequence.

The application of deuterium labeling and residual dipolar coupling constants in combination with other structural information has demonstrated the potential for significantly expanding the range of viable protein targets for structural analysis by NMR. A previous study by Clore et al. [(1999) J. Am. Chem. Soc. 121, 6513-6514] demonstrated that a significant improvement in the overall protein structure occurs with the combination of residual dipolar coupling constants and minimal tertiary long-range distance restraints. The analysis of NMR protein structures determined with minimal structural information is extended with a particular interest in the utility of these structures for a structure-based drug design program. As an example, the catalytic fragment of human fibroblast collagenase (MMP-1) was used to follow the effect of minimal restraint sets on the protein structure and its utility in drug design with a particular interest in the effect on the active site conformation. An MMP-1 structure that was calculated with the maximal number of restraints attainable with the constraint of a deuterated protein was shown to be very similar to a high-quality MMP-1 structure that was calculated from a complete set of restraints. The superposition of the active site backbone atoms for the high-quality and minimal restraint MMP-1 structures yielded an rmsd of 0.68 A where the size and shape of the S1' pocket are nearly identical. Additionally, an MMP-1-CGS-27023A complex based on a minimal set of NOE-based restraints reliably reproduced the structure of the complex, establishing the usefulness of the structures for drug design.