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A taste test for proteases

Nature Biotechnology volume 18, pages 149–150 (2000)Cite this article

The multitude of genome sequencing projects currently in progress will ensure the discovery of many genes for new enzymes in the coming decade1. A wealth of uncharacterized open reading frames, several thousand from bacterial or fungal species and up to a hundred thousand or more from the human genome, will be available soon2. So before us lies an unprecedented and daunting task—determining what the products of all these genes are doing in the cell. Biological function can be determined in numerous ways, but in the protease world, it is best defined by target sequence specificity3. In this issue, Backes et al.4 describe a new method for exploring this specificity small, focused combinatorial peptide libraries. They have developed a unique chemistry for peptide synthesis that, unlike previous methods, allows protease specificity to be defined rapidly not only by the amino acid just before the cleavage point, but also positions several amino acid residues removed.

Proteolytic enzymes usually contain an elongated active site cleft that allows 6–10 amino acids of the substrate to make contact with the binding site. For many enzymes, primary specificity is determined by contacts with the amino acid positioned just before the cleavage point (the P1 residue). The peptidase active site has a well-defined subsite pocket to snugly accommodate the side chain of residue P1. This anchors the substrate sequence in the active site so that the catalytic amino acids can act upon the scissile peptide bond (i.e., the one that marks the cleavage point). In some enzymes, only the P1 residue and the following amino acid (the P1′ residue) will contribute to interactions that define the cleavage point. However, in a large and growing group of peptidases, cleavage specificity is defined by interactions that take place along the entire active site cleft. Exploiting the extended interactions along the binding site may be essential for restricting cleavage to only one peptide bond of a particular protein, among all the other vulnerable peptide bonds within the cell.

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Figure 1: Schematic diagram of complex of protease with unfolded protein substrate in the binding groove.

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  1. distinguished professor of biochemistry and molecular biology, University of Florida College of Medicine, P.O. Box 100245, Gainesville, 32610-0245, FL

    Benn Dunn

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  1. Benn Dunn
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Dunn, B. A taste test for proteases. Nat Biotechnol 18, 149–150 (2000). https://doi.org/10.1038/72591

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