Structural Organization in the Serine Proteases

Abstract

We have been developing computational approaches to increase our ability to analyze the growing body of three-dimensional structural data with applications centered about the serine proteases. The emphasis of these approaches is to compare and contrast macromolecules at the separate levels of secondary, tertiary, and quaternary structure. Our assumption is that in functionally related molecules, regions of structural and/or physicochemical similarity will exhibit functional similarity; regions that are different in structure and/or physicochemical properties will function differently and, therefore, be the source of specificity. Based on this assumption, the independent observations from studies of these enzymes in solution and in biological systems are combined with the structural observations from X-ray crystallographic analysis. A goal of the present research effort is to probe the biomolecular architecture of the serine proteases to evaluate the role of the three-dimensional structure beyond that of the active site in determining recognition and reactivity determinants for these enzymes, and to determine those principles that might be applied successfully to other enzyme systems as well. Of particular note has been our observation of a macromolecular recognition surface which occurs as a topographical feature outside of the active site and under evolutionary control to produce specificity towards macromolecular substrates and inhibitors. In additon we have established the important role of conformational changes that occur beyond the active site of an enzyme and differentiate between natural and synthetic inhibitor-enzyme interactions. This suggests that the specificity and reactivity determinants of a macromolecule are derived from its architecture and structural organization.

Comparison of various molecular forms of bovine trypsin: correlation of infrared spectra with x-ray crystal structures

“…However, the inherent complexity, size, and lack of symmetry in globular proteins often prevent direct application of these results to protein spectra. Aided by tools developed for macromolecular structure analysis [e.g., see Levitt and Greer (1977), Kabsch and Sander (1983), and Liebman (1986)], comparisons of IR spectra with high-resolution crystallographically determined protein structures can establish necessary spectra-structure correlations Arrondo et al, 1988) as well as verify previous assignments which have often been based simply on empirical observations.…”

“…The atomic resolution of a number of these structures is high (1.5-1.8 Á for proteins used in this study). Liebman (1986) has reported an extensive analysis of these structures using tools developed for analysis of secondary and tertiary structure in proteins as well as methods for discerning small conformational differences in closely related molecules such as these. This study reported that the conformational changes which occur upon the transition between the different molecular states of bovine trypsin are localized within well-defined regions of the molecule and, thus, the overall backbone conformation remains unchanged.…”

Inhibitor-Induced Structural Changes in Serine Proteases Monitored by Fourier Transform Infrared Spectroscopy

“…Based on the difference LD plot, the main difference between the two structures is in the region between residues 168-174 (positions are referred to based on the ordered residue list in β-trypsin, i.e. 1-223 for 223 residues in β-trypsin (20). The seventh residue in trypsinogen, isoleucine, assumes position number 1.).…”

“…All computational methods used for analysis of crystallographic structure data are based on algorithms of Liebman and co-workers (19)(20)(21)(22).…”

“…To compare two forms of the same protein, their respective LD's are subtracted pairwise and a plot will highlight regions that undergo change (20). The BDA is a dihedral angle between two planes, one containing i, i+1, i+2 and the second containing i+1, i+2, 1+3.…”