Quantitative characterization of furin specificity. Energetics of substrate discrimination using an internally consistent set of hexapeptidyl methylcoumarinamides

Damian J Krysan; Nathan C Rockwell; Robert S Fuller

doi:10.1074/jbc.274.33.23229

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Quantitative characterization of furin specificity. Energetics of substrate discrimination using an internally consistent set of hexapeptidyl methylcoumarinamides

Journal article

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Quantitative characterization of furin specificity. Energetics of substrate discrimination using an internally consistent set of hexapeptidyl methylcoumarinamides

Damian J Krysan, Nathan C Rockwell and Robert S Fuller

The Journal of biological chemistry, Vol.274(33), pp.23229-23234

08/13/1999

DOI: 10.1074/jbc.274.33.23229

PMID: 10438496

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Abstract

Furin, an essential mammalian proprotein processing enzyme of the kexin/furin family of subtilisin-related eukaryotic processing proteases, is implicated in maturation of substrates involved in development, signaling, coagulation, and pathogenesis. We examined the energetics of furin specificity using a series of peptidyl methylcoumarinamide substrates. In contrast to previous reports, we found that furin can cleave such substrates with kinetics comparable to those observed with extended peptides and physiological substrates. With the best of these hexapeptidyl methylcoumarinamides, furin displayed k(cat)/K(m) values greater than 10(6) M(-1) s(-1). Furin exhibited striking substrate inhibition with hexapeptide but not tetrapeptide substrates, an observation of significance to the evaluation of peptide-based furin inhibitors. Quantitative comparison of furin and Kex2 recognition at P(1), P(2), and P(4) demonstrates that whereas interactions at P(1) make comparable contributions to catalysis by the two enzymes, furin exhibited a approximately 10-fold lesser dependence on P(2) recognition but a 10-100-fold greater dependence on P(4) recognition. Furin has recently been shown to exhibit P(6) recognition and we found that this interaction contributes approximately 1.4 kcal/mol toward catalysis independent of the nature of the P(4) residue. We have also shown that favorable residues at P(2) and P(6) will compensate for less than optimal residues at either P(1) or P(4). The quantitative analysis of furin and Kex2 specificity sharply distinguish the nature of substrate recognition by the processing and degradative members of subtilisin-related proteases.

Energy Metabolism

Amino Acid Sequence

Coumarins - metabolism

Molecular Sequence Data

Substrate Specificity

Amides

Subtilisins - antagonists & inhibitors

Hydrolysis

Saccharomyces cerevisiae Proteins

Furin

Subtilisins - metabolism

Kinetics

Proprotein Convertases

Serine Proteinase Inhibitors - pharmacology

Details

Title: Subtitle: Quantitative characterization of furin specificity. Energetics of substrate discrimination using an internally consistent set of hexapeptidyl methylcoumarinamides
Creators: Damian J Krysan - Department of Biological Chemistry, University of Michigan Medical School, Ann Arbor, Michigan 48109, USA
Nathan C Rockwell
Robert S Fuller
Resource Type: Journal article
Publication Details: The Journal of biological chemistry, Vol.274(33), pp.23229-23234
DOI: 10.1074/jbc.274.33.23229
PMID: 10438496
NLM abbreviation: J Biol Chem
ISSN: 0021-9258
eISSN: 1083-351X
Language: English
Date published: 08/13/1999
Academic Unit: Microbiology and Immunology; Stead Family Department of Pediatrics; Infectious Disease (Pediatrics)
Record Identifier: 9984093475502771

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