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Chem Biol. 2015 Aug 20;22(8):995-1001. doi: 10.1016/j.chembiol.2015.07.010. Epub 2015 Aug 6.

Design and Synthesis of Activity-Based Probes and Inhibitors for Bleomycin Hydrolase.

Author information

1
Departments of Pathology and Microbiology and Immunology, Stanford University School of Medicine, 300 Pasteur Drive, Stanford, CA 94305, USA.
2
Division of Bioorganic Chemistry, Faculty of Chemistry, Wroclaw University of Technology, Wyb. Wyspianskiego 27, 50-370 Wroclaw, Poland.
3
Departments of Pathology and Microbiology and Immunology, Stanford University School of Medicine, 300 Pasteur Drive, Stanford, CA 94305, USA. Electronic address: mbogyo@stanford.edu.

Abstract

Bleomycin hydrolase (BLMH) is a neutral cysteine aminopeptidase that has been ascribed roles in many physiological and pathological processes, yet its primary biological function remains enigmatic. In this work, we describe the results of screening of a library of fluorogenic substrates to identify non-natural amino acids that are optimally recognized by BLMH. This screen identified several substrates with kcat/KM values that are substantially improved over the previously reported fluorogenic substrates for this enzyme. The substrate sequences were used to design activity-based probes that showed potent labeling of recombinant BLMH as well as endogenously expressed BLMH in cell extracts, and in intact cells. Importantly, we identify potent BLMH inhibitors that are able to fully inhibit endogenous BLMH activity in intact cells. These probes and inhibitors will be valuable new reagents to study BLMH function in cellular and animal models of human diseases where BLMH is likely to be involved.

PMID:
26256478
PMCID:
PMC4546515
DOI:
10.1016/j.chembiol.2015.07.010
[Indexed for MEDLINE]
Free PMC Article

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