Structural engineering of the HIV‐1 protease molecule with a β‐turn mimic of fixed geometry

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Abstract
An important goal in the de novo design of enzymes is the control of molecular geometry. To this end, an analog of the protease from human immunodeficiency virus 1 (HIV-1 protease) was prepared by total chemical synthesis, containing a constrained, nonpeptidic type II' β-turn mimic of predetermined three-dimensional structure. The mimic β-turn replaced residues Gly16, 17 in each subunit of the homodimeric molecule. These residues constitute the central amino acids of two symmetry-related type I' β-turns in the native, unliganded enzyme. The β-turn mimic-containing enzyme analog was fully active, possessed the same substrate specificity as the Gly16, 17-containing enzyme, and showed enhanced resistance to thermal inactivation. These results indicate that the precise geometry of the β-turn at residues 15–18 in each subunit is not critical for activity, and that replacement of the native sequence with a rigid β-turn mimic can lead to enhanced protein stability. Finally, the successful incorporation of a fixed element of secondary structure illustrates the potential of a “molecular kit set” approach to protein design and synthesis.