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Title: Functional Characterization of Four Putative δ1-Pyrroline-5-Carboxylate Reductases from Bacillus subtilis

Abstract

In most living organisms, the amino acid proline is synthesized starting from both glutamate and ornithine. In prokaryotes, in the absence of an ornithine cyclodeaminase that has been identified to date only in a small number of soil and plant bacteria, these pathways share the last step, the reduction of delta(1)-pyrroline-5-carboxylate (P5C) catalyzed by P5C reductase (EC 1.5.1.2). In several species, multiple forms of P5C reductase have been reported, possibly reflecting the dual function of proline. Aside from its common role as a building block of proteins, proline is indeed also involved in the cellular response to osmotic and oxidative stress conditions. Genome analysis of Bacillus subtilis identifies the presence of four genes (ProH, ProI, ProG, and ComER) that, based on bioinformatic and phylogenic studies, were defined as respectively coding a putative P5C reductase. Here we describe the cloning, heterologous expression, functional analysis and small-angle X-ray scattering studies of the four affinity-purified proteins. Results showed that two of them, namely ProI and ComER, lost their catalytic efficiency or underwent subfunctionalization. In the case of ComER, this could be likely explained by the loss of the ability to form a dimer, which has been previously shown to be an essential structuralmore » feature of the catalytically active P5C reductase. The properties of the two active enzymes are consistent with a constitutive role for ProG, and suggest that ProH expression may be beneficial to satisfy an increased need for proline.« less

Authors:
; ; ;
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States)
Sponsoring Org.:
National Institutes of Health (NIH) - National Institute of General Medical Sciences; National Institutes of Health (NIH) - National Institute of Allergy and Infectious Diseases (NIAID); University of Ferrara
OSTI Identifier:
1393852
DOE Contract Number:  
AC02-06CH11357
Resource Type:
Journal Article
Journal Name:
Frontiers in Microbiology
Additional Journal Information:
Journal Volume: 8; Journal ID: ISSN 1664-302X
Publisher:
Frontiers Research Foundation
Country of Publication:
United States
Language:
English
Subject:
isoenzyme properties; Bacillus subtilis; P5C reductase; oligomeric structure; product inhibition; proline synthesis; substrate ambiguity

Citation Formats

Forlani, Giuseppe, Nocek, Boguslaw, Chakravarthy, Srinivas, and Joachimiak, Andrzej. Functional Characterization of Four Putative δ1-Pyrroline-5-Carboxylate Reductases from Bacillus subtilis. United States: N. p., 2017. Web. doi:10.3389/fmicb.2017.01442.
Forlani, Giuseppe, Nocek, Boguslaw, Chakravarthy, Srinivas, & Joachimiak, Andrzej. Functional Characterization of Four Putative δ1-Pyrroline-5-Carboxylate Reductases from Bacillus subtilis. United States. doi:10.3389/fmicb.2017.01442.
Forlani, Giuseppe, Nocek, Boguslaw, Chakravarthy, Srinivas, and Joachimiak, Andrzej. Wed . "Functional Characterization of Four Putative δ1-Pyrroline-5-Carboxylate Reductases from Bacillus subtilis". United States. doi:10.3389/fmicb.2017.01442.
@article{osti_1393852,
title = {Functional Characterization of Four Putative δ1-Pyrroline-5-Carboxylate Reductases from Bacillus subtilis},
author = {Forlani, Giuseppe and Nocek, Boguslaw and Chakravarthy, Srinivas and Joachimiak, Andrzej},
abstractNote = {In most living organisms, the amino acid proline is synthesized starting from both glutamate and ornithine. In prokaryotes, in the absence of an ornithine cyclodeaminase that has been identified to date only in a small number of soil and plant bacteria, these pathways share the last step, the reduction of delta(1)-pyrroline-5-carboxylate (P5C) catalyzed by P5C reductase (EC 1.5.1.2). In several species, multiple forms of P5C reductase have been reported, possibly reflecting the dual function of proline. Aside from its common role as a building block of proteins, proline is indeed also involved in the cellular response to osmotic and oxidative stress conditions. Genome analysis of Bacillus subtilis identifies the presence of four genes (ProH, ProI, ProG, and ComER) that, based on bioinformatic and phylogenic studies, were defined as respectively coding a putative P5C reductase. Here we describe the cloning, heterologous expression, functional analysis and small-angle X-ray scattering studies of the four affinity-purified proteins. Results showed that two of them, namely ProI and ComER, lost their catalytic efficiency or underwent subfunctionalization. In the case of ComER, this could be likely explained by the loss of the ability to form a dimer, which has been previously shown to be an essential structural feature of the catalytically active P5C reductase. The properties of the two active enzymes are consistent with a constitutive role for ProG, and suggest that ProH expression may be beneficial to satisfy an increased need for proline.},
doi = {10.3389/fmicb.2017.01442},
journal = {Frontiers in Microbiology},
issn = {1664-302X},
number = ,
volume = 8,
place = {United States},
year = {2017},
month = {8}
}

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