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Article
Molecular Discrimination of Type-I over Type-II Methionyl Aminopeptidases
Biochemistry
  • Krzysztof Swierczek, Utah State University
  • Alicja J. Copik, Utah State University
  • Sabina I. Swierczek, Marquette University
  • Richard C. Holz, Marquette University
Document Type
Article
Publication Date
9-13-2005
Disciplines
Abstract
Two residues that are conserved in type-I methionyl aminopeptidases (MetAPs) but are absent in all type-II MetAPs are the cysteine residues (Escherichia coli MetAP-I:   C59 and C70) that reside at the back of the substrate recognition pocket. These Cys residues are 4.4 Å apart and do not form a disulfide bond. Since bacteria and fungi contain only type-I MetAPs while all human cells contain both type-I and type-II MetAPs, type-I MetAPs represent a novel antibiotic/antifungal target if type-I MetAPs can be specifically targeted over type-II. Based on reaction of the thiol-specific binding reagent 5,5‘-dithio-bis(2-nitrobenzoic acid) (DTNB) with the type-I MetAP from E. coli and the type-II MetAP from Pyrococcus furiosus, the type-I MetAP can be selectively inhibited. Verification that DTNB covalently binds to C59 in EcMetAP-I was obtained by mass spectrometry (MS) from reaction of DTNB with the C59A and C70A mutant EcMetAP-I enzymes. In addition, two inhibitors of EcMetAP-I, 5-iodopentaphosphonic acid (1) and 6-phosphonohexanoic acid (2), were designed and synthesized. The first was designed as a selective-C59 binding reagent while the second was designed as a simple competitive inhibitor of EcMetAP. Indeed, inhibitor 1 forms a covalent interaction with C59 based on activity assays and MS measurements, while 2 does not. These data indicate that type-I MetAPs can be selectively targeted over type-II MetAPs, suggesting that type-I MetAPs represent a new enzymatic target for antibacterial or antifungal agents.
Comments

BIochemistry, Vol. 44, No. 36 (September 13, 2005): 12049-12056. DOI.

Richard C. Holz was affiliated with Utah State University at the time of publication.

Citation Information
Krzysztof Swierczek, Alicja J. Copik, Sabina I. Swierczek and Richard C. Holz. "Molecular Discrimination of Type-I over Type-II Methionyl Aminopeptidases" Biochemistry (2005) ISSN: 0006-2960
Available at: http://works.bepress.com/richard_holz/17/