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The Neisseria gonorrhoeae Obg protein is an essential ribosome-associated GTPase and a potential drug target.

Zielke RA, Wierzbicki IH, Baarda BI, Sikora AE - BMC Microbiol. (2015)

Bottom Line: Serious complications associated with these infections are frequent and include pelvic inflammatory disease, ectopic pregnancy, and infertility.The cellular levels of Obg reach a maximum in the early logarithmic phase and remain constant throughout bacterial growth.Characterization of the GC Obg at the molecular and functional levels presented herein may facilitate the future targeting of this protein with small molecule inhibitors and the evaluation of identified lead compounds for bactericidal activity against GC and other drug-resistant bacteria.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmaceutical Sciences, College of Pharmacy, Oregon State University, 433 Weniger Hall, 103 SW Memorial Pl, Corvallis, OR, 97330, USA. Ryszard.Zielke@oregonstate.edu.

ABSTRACT

Background: Neisseria gonorrhoeae (GC) is a Gram-negative pathogen that most commonly infects mucosal surfaces, causing sexually transmitted urethritis in men and endocervicitis in women. Serious complications associated with these infections are frequent and include pelvic inflammatory disease, ectopic pregnancy, and infertility. The incidence of gonorrhea cases remains high globally while antibiotic treatment options, the sole counter measures against gonorrhea, are declining due to the remarkable ability of GC to acquire resistance. Evaluating of potential drug targets is essential to provide opportunities for developing antimicrobials with new mechanisms of action. We propose the GC Obg protein, belonging to the Obg/CgtA GTPase subfamily, as a potential target for the development of therapeutic interventions against gonorrhea, and in this study perform its initial functional and biochemical characterization.

Results: We report that NGO1990 encodes Obg protein, which is an essential factor for GC viability, associates predominantly with the large 50S ribosomal subunit, and is stably expressed under conditions relevant to infection of the human host. The anti-Obg antisera cross-reacts with a panel of contemporary GC clinical isolates, demonstrating the ubiquitous nature of Obg. The cellular levels of Obg reach a maximum in the early logarithmic phase and remain constant throughout bacterial growth. The in vitro binding and hydrolysis of the fluorescent guanine nucleotide analogs mant-GTP and mant-GDP by recombinant wild type and T192AT193A mutated variants of Obg are also assessed.

Conclusions: Characterization of the GC Obg at the molecular and functional levels presented herein may facilitate the future targeting of this protein with small molecule inhibitors and the evaluation of identified lead compounds for bactericidal activity against GC and other drug-resistant bacteria.

No MeSH data available.


Related in: MedlinePlus

ObgGC domain architecture. The individual structural domains of ObgGC are shown in green. The N-terminal domain (amino acids 3–158) is glycine-rich. The central, GTP-binding domain (residues 160–348) includes two switch elements (switch I and switch II) and five conserved G motifs (G1-G5; indicated in blue boxes). The C-terminal domain contains clusters of acidic residues. The conserved T192 and T193 residues within the G2 motifs and introduced substitutions are designated in red
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Fig1: ObgGC domain architecture. The individual structural domains of ObgGC are shown in green. The N-terminal domain (amino acids 3–158) is glycine-rich. The central, GTP-binding domain (residues 160–348) includes two switch elements (switch I and switch II) and five conserved G motifs (G1-G5; indicated in blue boxes). The C-terminal domain contains clusters of acidic residues. The conserved T192 and T193 residues within the G2 motifs and introduced substitutions are designated in red

Mentions: Analysis of the predicted amino acid sequence of NGO1990 from FA1090 revealed a typical structure of Obg GTPases (Fig. 1) and significant similarities to other Obg proteins (Table 1). The N-terminal domain of ObgGC (amino acids 3–158) contains 26 glycine residues, similar to the B. subtilis Obg [20]. The central, GTP-binding domain (residues 160–348), includes five conserved G motifs (G1-G5) and two switch elements (switch I and switch II) that determine the active or inactive state of the G protein [15, 16]. As expected, the C-terminal domain shows the lowest conservation when compared to Obg homologs from different bacterial species (Additional file 1: Figure S1). Nevertheless, this region of ObgGC contains clusters of acidic residues, a feature characteristic of Obg proteins [17, 33]. This charged C-terminus has been shown to be important for Obg association with 50S ribosomal particles in C. crescentus as well as GTP and GDP binding in V. harveyi [17, 26].Fig. 1


The Neisseria gonorrhoeae Obg protein is an essential ribosome-associated GTPase and a potential drug target.

Zielke RA, Wierzbicki IH, Baarda BI, Sikora AE - BMC Microbiol. (2015)

ObgGC domain architecture. The individual structural domains of ObgGC are shown in green. The N-terminal domain (amino acids 3–158) is glycine-rich. The central, GTP-binding domain (residues 160–348) includes two switch elements (switch I and switch II) and five conserved G motifs (G1-G5; indicated in blue boxes). The C-terminal domain contains clusters of acidic residues. The conserved T192 and T193 residues within the G2 motifs and introduced substitutions are designated in red
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4487204&req=5

Fig1: ObgGC domain architecture. The individual structural domains of ObgGC are shown in green. The N-terminal domain (amino acids 3–158) is glycine-rich. The central, GTP-binding domain (residues 160–348) includes two switch elements (switch I and switch II) and five conserved G motifs (G1-G5; indicated in blue boxes). The C-terminal domain contains clusters of acidic residues. The conserved T192 and T193 residues within the G2 motifs and introduced substitutions are designated in red
Mentions: Analysis of the predicted amino acid sequence of NGO1990 from FA1090 revealed a typical structure of Obg GTPases (Fig. 1) and significant similarities to other Obg proteins (Table 1). The N-terminal domain of ObgGC (amino acids 3–158) contains 26 glycine residues, similar to the B. subtilis Obg [20]. The central, GTP-binding domain (residues 160–348), includes five conserved G motifs (G1-G5) and two switch elements (switch I and switch II) that determine the active or inactive state of the G protein [15, 16]. As expected, the C-terminal domain shows the lowest conservation when compared to Obg homologs from different bacterial species (Additional file 1: Figure S1). Nevertheless, this region of ObgGC contains clusters of acidic residues, a feature characteristic of Obg proteins [17, 33]. This charged C-terminus has been shown to be important for Obg association with 50S ribosomal particles in C. crescentus as well as GTP and GDP binding in V. harveyi [17, 26].Fig. 1

Bottom Line: Serious complications associated with these infections are frequent and include pelvic inflammatory disease, ectopic pregnancy, and infertility.The cellular levels of Obg reach a maximum in the early logarithmic phase and remain constant throughout bacterial growth.Characterization of the GC Obg at the molecular and functional levels presented herein may facilitate the future targeting of this protein with small molecule inhibitors and the evaluation of identified lead compounds for bactericidal activity against GC and other drug-resistant bacteria.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmaceutical Sciences, College of Pharmacy, Oregon State University, 433 Weniger Hall, 103 SW Memorial Pl, Corvallis, OR, 97330, USA. Ryszard.Zielke@oregonstate.edu.

ABSTRACT

Background: Neisseria gonorrhoeae (GC) is a Gram-negative pathogen that most commonly infects mucosal surfaces, causing sexually transmitted urethritis in men and endocervicitis in women. Serious complications associated with these infections are frequent and include pelvic inflammatory disease, ectopic pregnancy, and infertility. The incidence of gonorrhea cases remains high globally while antibiotic treatment options, the sole counter measures against gonorrhea, are declining due to the remarkable ability of GC to acquire resistance. Evaluating of potential drug targets is essential to provide opportunities for developing antimicrobials with new mechanisms of action. We propose the GC Obg protein, belonging to the Obg/CgtA GTPase subfamily, as a potential target for the development of therapeutic interventions against gonorrhea, and in this study perform its initial functional and biochemical characterization.

Results: We report that NGO1990 encodes Obg protein, which is an essential factor for GC viability, associates predominantly with the large 50S ribosomal subunit, and is stably expressed under conditions relevant to infection of the human host. The anti-Obg antisera cross-reacts with a panel of contemporary GC clinical isolates, demonstrating the ubiquitous nature of Obg. The cellular levels of Obg reach a maximum in the early logarithmic phase and remain constant throughout bacterial growth. The in vitro binding and hydrolysis of the fluorescent guanine nucleotide analogs mant-GTP and mant-GDP by recombinant wild type and T192AT193A mutated variants of Obg are also assessed.

Conclusions: Characterization of the GC Obg at the molecular and functional levels presented herein may facilitate the future targeting of this protein with small molecule inhibitors and the evaluation of identified lead compounds for bactericidal activity against GC and other drug-resistant bacteria.

No MeSH data available.


Related in: MedlinePlus