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Title: Kinetic characterization of Vibrio cholerae ApbE: Substrate specificity and regulatory mechanisms

Abstract

ApbE is a member of a novel family of flavin transferases that incorporates flavin mononucleotide (FMN) to subunits of diverse respiratory complexes, which fulfill important homeostatic functions. In this work a detailed characterization of Vibrio cholerae ApbE physiologic activity, substrate specificity and pH dependency was carried out. The data obtained show novel characteristics of the regulation and function of this family. For instance, our experiments indicate that divalent cations are essential for ApbE function, and that the selectivity depends largely on size and the coordination sphere of the cation. Our data also show that ApbE regulation by pH, ADP and potassium is an important mechanism that enhances the adaptation, survival and colonization of V. cholerae in the small intestine. Moreover, studies of the pH-dependency of the activity show that the reaction is favored under alkaline conditions, with a pKa of 8.4. These studies, together with sequence and structure analysis allowed us to identify His257, which is absolutely conserved in the family, as a candidate for the residue whose deprotonation controls the activity. Remarkably, the mutant H257G abolished the flavin transfer activity, strongly indicating that this residue plays an important role in the catalytic mechanism of ApbE.

Authors:
 [1];  [1];  [1];  [1];  [2];  [1]; ORCiD logo [1]
  1. Illinois Inst. of Technology, Chicago, IL (United States)
  2. Illinois Inst. of Technology, Chicago, IL (United States); Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Org.:
Illinois Institute of Technology
OSTI Identifier:
1432860
Resource Type:
Accepted Manuscript
Journal Name:
PLoS ONE
Additional Journal Information:
[ Journal Volume: 12; Journal Issue: 10]; Journal ID: ISSN 1932-6203
Publisher:
Public Library of Science
Country of Publication:
United States
Language:
ENGLISH
Subject:
59 BASIC BIOLOGICAL SCIENCES; Flavin; Vibrio cholerae; Cations; Cofactors (biochemistry); Enzyme regulation; Ultraviolet-visible; Spectroscopy; Hydrolysis; Phosphates

Citation Formats

Fang, Xuan, Liang, Pingdong, Raba, Daniel Alexander, Rosas-Lemus, Mónica, Chakravarthy, Srinivas, Tuz, Karina, and Juárez, Oscar. Kinetic characterization of Vibrio cholerae ApbE: Substrate specificity and regulatory mechanisms. United States: N. p., 2017. Web. doi:10.1371/journal.pone.0186805.
Fang, Xuan, Liang, Pingdong, Raba, Daniel Alexander, Rosas-Lemus, Mónica, Chakravarthy, Srinivas, Tuz, Karina, & Juárez, Oscar. Kinetic characterization of Vibrio cholerae ApbE: Substrate specificity and regulatory mechanisms. United States. doi:10.1371/journal.pone.0186805.
Fang, Xuan, Liang, Pingdong, Raba, Daniel Alexander, Rosas-Lemus, Mónica, Chakravarthy, Srinivas, Tuz, Karina, and Juárez, Oscar. Tue . "Kinetic characterization of Vibrio cholerae ApbE: Substrate specificity and regulatory mechanisms". United States. doi:10.1371/journal.pone.0186805. https://www.osti.gov/servlets/purl/1432860.
@article{osti_1432860,
title = {Kinetic characterization of Vibrio cholerae ApbE: Substrate specificity and regulatory mechanisms},
author = {Fang, Xuan and Liang, Pingdong and Raba, Daniel Alexander and Rosas-Lemus, Mónica and Chakravarthy, Srinivas and Tuz, Karina and Juárez, Oscar},
abstractNote = {ApbE is a member of a novel family of flavin transferases that incorporates flavin mononucleotide (FMN) to subunits of diverse respiratory complexes, which fulfill important homeostatic functions. In this work a detailed characterization of Vibrio cholerae ApbE physiologic activity, substrate specificity and pH dependency was carried out. The data obtained show novel characteristics of the regulation and function of this family. For instance, our experiments indicate that divalent cations are essential for ApbE function, and that the selectivity depends largely on size and the coordination sphere of the cation. Our data also show that ApbE regulation by pH, ADP and potassium is an important mechanism that enhances the adaptation, survival and colonization of V. cholerae in the small intestine. Moreover, studies of the pH-dependency of the activity show that the reaction is favored under alkaline conditions, with a pKa of 8.4. These studies, together with sequence and structure analysis allowed us to identify His257, which is absolutely conserved in the family, as a candidate for the residue whose deprotonation controls the activity. Remarkably, the mutant H257G abolished the flavin transfer activity, strongly indicating that this residue plays an important role in the catalytic mechanism of ApbE.},
doi = {10.1371/journal.pone.0186805},
journal = {PLoS ONE},
number = [10],
volume = [12],
place = {United States},
year = {2017},
month = {10}
}

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