The Escherichia coli NadR Regulator Is Endowed with Nicotinamide Mononucleotide Adenylyltransferase Activity

Abstract

The first identification and characterization of a catalytic activity associated with NadR protein is reported. A computer-aided search for sequence similarity revealed the presence in NadR of a 29-residue region highly conserved among known nicotinamide mononucleotide adenylyltransferases. The Escherichia coli nadR gene was cloned into a T7-based vector and overexpressed. In addition to functionally specific DNA binding properties, the homogeneous recombinant protein catalyzes NAD synthesis from nicotinamide mononucleotide and ATP.

Comparative genomics of metabolic pathways inMycobacteriumspecies: gene duplication, gene decay and lateral gene transfer

“…The virS gene encoding a bacterial virulence-regulating protein is also absent in M. smegmatis and M. avium. The absence of virS further confirms its role as a regulator of genes that differentiate the M. tuberculosis complex from other mycobacterial species (Raffaelli et al, 1999), whereas the possible effects of the absence of nadR, a gene encoding a bifunctional protein that has a role in the transport of nicotinamide mononucleotide, remain to be tested. Moreover, the nadR homolog in M. tuberculosis and M. bovis is shorter (323 amino acids) than the corresponding gene in Escherichia coli (417 amino acids), losing a portion of the N-terminal region, indicating a lack of repressor function (Hill et al, 1998).…”

Metabolic engineering ofEscherichia colifor optimized biosynthesis of nicotinamide mononucleotide, a noncanonical redox cofactor

“…When 200 μM NMN + was directly fed to the cells expressing an NMN + transporter from S. enterica PnuC* [29] (on a multiple-copy plasmid pWB302), growth was restored to levels comparable to wild type cells after 6 hours ( Figure 2A). In contrast, cells carrying a control plasmid (pWB301) could not grow, suggesting the basal level of NMN + in E. coli cells does not cause background issues in this growth-based screening, possibly because E. coli NadR has low affinity towards NMN + [30]. These results demonstrate that the screening platform is functioning properly, and cell growth is serving as a readout for intracellular NMN + level.…”

“…Moreover, both efforts did not study the physiological response in E. coli to NMN + accumulation. Beside functioning as a cofactor, NAD + is also a universal signaling compound that allosterically controls key enzymes and transcriptional regulators in response to the fluctuating cellular redox state [30,[33][34][35][36]. Since NMN + is an analog of NAD + , it is an open question whether NMN + , when it accumulates to a high level, can also interact with the numerous proteins that are modulated by NAD + .…”

“…NAD + has been suggested to allosterically modulate NadR's function [32]. Given NadR's capability to also recognize NMN + [30], further studies are needed to investigate whether NMN + binding induces conformational change in the DNA-binding domain of NadR and modulates its function as a transcriptional regulator.…”

Structural and Functional Characterization of NadR from Lactococcus lactis

“…Structurally the NMNAT domain is similar to archaeal NMNAT, whereas the RNK domain is similar to yeast thymidylate kinase [22]. For the NadR gene of E. coli, it has been shown experimentally that the NMNAT domain is indeed responsible for the transfer of the AMP moiety from ATP to NMN [23]. A crystal structure of the E. coli NMNAT domain of NadR with NAD bound has been reported [24].…”