No abstract

The NMR chemical shifts of certain atomic nuclei in proteins ( 1 H␣, 13 C␣, and 13 C␤) depend sensitively on whether or not the amino acid residue is part of a secondary structure (␣-helix, ␤-sheet), and if so, whether it is helix or sheet. The physical origin of the different chemical shifts of atomic nuclei in ␣-helices versus ␤-sheets is a problem of long standing. We report that the chemical shift contributions arising from secondary structure (secondary structure shifts) depend strongly on the extent of exposure to solvent. This behavior is observed for 1 H␣, 13 C␣, and 13 C␤ (sheet), but not for 13 C␤ (helix), whose secondary structure shifts are small. When random coil values are subtracted from the chemical shifts of all 1 H␣ nuclei (Pro residues excluded) and the residual chemical shifts are summed to plot the mean values against solvent exposure, the results give a funnel-shaped curve that approaches a small value at full-solvent exposure. When chemical shifts are plotted instead against E local, the electrostatic contribution to conformational energy produced by local dipole-dipole interactions, a well characterized dependence of 1 H␣ chemical shifts on Elocal is found. The slope of this plot varies with both the type of amino acid and the extent of solvent exposure. These results indicate that secondary structure shifts are produced chiefly by the electric field of the protein, which is screened by water dipoles at residues in contact with solvent.protein solvation ͉ secondary structure shifts T he chemical shift index method (1, 2) is commonly used to assign protein secondary structures. This method is based on the secondary structure shift, which is the difference between the observed chemical shift and the random coil value assigned to this amino acid type in the unfolded conformation. Assignment of secondary structure is a useful intermediate step in determining the 3D solution structure of a protein from NMR data. Secondary structure assignment is also useful in predicting the 3D structure of a protein from its amino acid sequence. A threading method based on chemical shift data has been developed (2) for testing whether an amino acid sequence can fold to form a given 3D structure. Understanding the origin of the distinctive secondary structure shifts of helices and sheets is a problem of considerable interest.Protein chemical shifts, for a given atomic nucleus and amino acid type, display large variations, and the factors that control these variations are poorly understood. The reason why the chemical shift index method works so well for assigning secondary structures is that the secondary structure shift has opposite signs for ␣-helix and ␤-sheet (1-3). Several proposals have been made (4-10) regarding the physical origin of the different chemical shifts found in ␣-helices and ␤-sheets. For both the 1 H ␣ and the 13 C ␣ nuclei, protein chemical shifts are said to depend chiefly on , torsion angles and on the random coil values (2). The protein electric field may contribute to chemical shifts (...

A wide variety of pathogens have acquired antimicrobial resistance as an inevitable evolutionary response to the extensive use of antibacterial agents. In particular, one of the most widely used antibiotic structural classes is the beta-lactams, in which the most common and the most efficient mechanism of bacterial resistance is the synthesis of beta-lactamases. Class C beta-lactamase enzymes are primarily cephalosporinases, mostly chromosomally encoded, and are inducible by exposure to some beta-lactam agents and resistant to inhibition by marketed beta-lactamase inhibitors. In an ongoing effort to alleviate this problem a series of novel 4-substituted trinems was designed and synthesized. Significant in vitro inhibitory activity was measured against the bacterial beta-lactamases of class C and additionally against class A. The lead compound LK-157 was shown to be a potent mechanism-based inactivator. Acylation of the active site Ser 64 of the class C enzyme beta-lactamase was observed in the solved crystal structures of two inhibitors complexes to AmpC enzyme from E. cloacae. Structure-activity relationships in the series reveal the importance of the trinem scaffold for inhibitory activity and the interesting potential of the series for further development.