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Title: Functional Proteomic And Structural Insights Into Molecular Recognition in the Nitrilase Family Enzymes

Abstract

Nitrilases are a large and diverse family of nonpeptidic C-N hydrolases. The mammalian genome encodes eight nitrilase enzymes, several of which remain poorly characterized. Prominent among these are nitrilase-1 (Nit1) and nitrilase-2 (Nit2), which, despite having been shown to exert effects on cell growth and possibly serving as tumor suppressor genes, are without known substrates or selective inhibitors. In previous studies, we identified several nitrilases, including Nit1 and Nit2, as targets for dipeptide-chloroacetamide activity-based proteomics probes. Here, we have used these probes, in combination with high-resolution crystallography and molecular modeling, to systematically map the active site of Nit2 and identify residues involved in molecular recognition. We report the 1.4 {angstrom} crystal structure of mouse Nit2 and use this structure to identify residues that discriminate probe labeling between the Nit1 and Nit2 enzymes. Interestingly, some of these residues are conserved across all vertebrate Nit2 enzymes and, conversely, not found in any vertebrate Nit1 enzymes, suggesting that they are key discriminators of molecular recognition between these otherwise highly homologous enzymes. Our findings thus point to a limited set of active site residues that establish distinct patterns of molecular recognition among nitrilases and provide chemical probes to selectively perturb the function of thesemore » enzymes in biological systems.« less

Authors:
; ; ; ; ; ; ;
Publication Date:
Research Org.:
Stanford Linear Accelerator Center (SLAC)
Sponsoring Org.:
USDOE
OSTI Identifier:
958628
Report Number(s):
SLAC-REPRINT-2009-061
Journal ID: ISSN 0006-2960; BICHAW; TRN: US201001%%776
DOE Contract Number:
AC02-76SF00515
Resource Type:
Journal Article
Resource Relation:
Journal Name: Biochem. 47:13514,2008; Journal Volume: 47; Journal Issue: 51
Country of Publication:
United States
Language:
English
Subject:
36 MATERIALS SCIENCE; CRYSTAL STRUCTURE; CRYSTALLOGRAPHY; DISCRIMINATORS; ENZYMES; FUNCTIONALS; GENES; HYDROLASES; NEOPLASMS; PROBES; RESIDUES; SUBSTRATES; TARGETS; VERTEBRATES; Other,CHEM, BIO

Citation Formats

Barglow, K.T., Saikatendu, K., Bracey, M.H., Huey, R., Morris, G.M., Olson, A.J., Stevens, R.C., and Cravatt, B.F.. Functional Proteomic And Structural Insights Into Molecular Recognition in the Nitrilase Family Enzymes. United States: N. p., 2009. Web.
Barglow, K.T., Saikatendu, K., Bracey, M.H., Huey, R., Morris, G.M., Olson, A.J., Stevens, R.C., & Cravatt, B.F.. Functional Proteomic And Structural Insights Into Molecular Recognition in the Nitrilase Family Enzymes. United States.
Barglow, K.T., Saikatendu, K., Bracey, M.H., Huey, R., Morris, G.M., Olson, A.J., Stevens, R.C., and Cravatt, B.F.. Mon . "Functional Proteomic And Structural Insights Into Molecular Recognition in the Nitrilase Family Enzymes". United States. doi:.
@article{osti_958628,
title = {Functional Proteomic And Structural Insights Into Molecular Recognition in the Nitrilase Family Enzymes},
author = {Barglow, K.T. and Saikatendu, K. and Bracey, M.H. and Huey, R. and Morris, G.M. and Olson, A.J. and Stevens, R.C. and Cravatt, B.F.},
abstractNote = {Nitrilases are a large and diverse family of nonpeptidic C-N hydrolases. The mammalian genome encodes eight nitrilase enzymes, several of which remain poorly characterized. Prominent among these are nitrilase-1 (Nit1) and nitrilase-2 (Nit2), which, despite having been shown to exert effects on cell growth and possibly serving as tumor suppressor genes, are without known substrates or selective inhibitors. In previous studies, we identified several nitrilases, including Nit1 and Nit2, as targets for dipeptide-chloroacetamide activity-based proteomics probes. Here, we have used these probes, in combination with high-resolution crystallography and molecular modeling, to systematically map the active site of Nit2 and identify residues involved in molecular recognition. We report the 1.4 {angstrom} crystal structure of mouse Nit2 and use this structure to identify residues that discriminate probe labeling between the Nit1 and Nit2 enzymes. Interestingly, some of these residues are conserved across all vertebrate Nit2 enzymes and, conversely, not found in any vertebrate Nit1 enzymes, suggesting that they are key discriminators of molecular recognition between these otherwise highly homologous enzymes. Our findings thus point to a limited set of active site residues that establish distinct patterns of molecular recognition among nitrilases and provide chemical probes to selectively perturb the function of these enzymes in biological systems.},
doi = {},
journal = {Biochem. 47:13514,2008},
number = 51,
volume = 47,
place = {United States},
year = {Mon May 11 00:00:00 EDT 2009},
month = {Mon May 11 00:00:00 EDT 2009}
}
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