Pyrroloquinoline quinone (PQQ) is a small, redox-active molecule that serves as a cofactor for several bacterial dehydrogenases, introducing pathways for carbon utilization that confer a growth advantage. Early studies had implicated a ribosomally translated peptide as the substrate for PQQ production. This study presents a sequence and structure based analysis of the components of the pqq operon. We find the necessary components for PQQ production are present in 126 prokaryotes, most of which are Gram- negative and a number of which are pathogens. A total of five gene products, PqqA, PqqB, PqqC, PqqD and PqqE, are concluded to be obligatory for PQQ production. Three of the gene products in the pqq operon, PqqB, PqqC and PqqE, are members of large protein superfamilies. By combining evolutionary conservation patterns with information from three-dimensional structures, we are able to differentiate the gene products involved in PQQ biosynthesis from those with divergent functions. The observed persistence of a conserved gene order within analyzed operons strongly suggests a role for protein/protein interactions in the course of cofactor biosynthesis. These studies propose previously unidentified roles for several of the gene products as well as possible new targets for antibiotic design and application.
Traditional Chinese Medicines (TCMs) have been historically used to treat bacterial infections. However, the molecules responsible for these anti-infective properties and their potential mechanisms of action have remained elusive. Using a high-throughput assay for type III protein secretion in Salmonella enterica serovar Typhimurium, we discovered that several TCMs can attenuate this key virulence pathway without affecting bacterial growth. Amongst the active TCMs, we discovered that baicalein, a specific flavonoid from Scutellaria baicalensis, targets S. Typhimurium pathogenicity island-1 (SPI-1) type III secretion system (T3SS) effectors and translocases to inhibit bacterial invasion of epithelial cells. Structurally related flavonoids present in other TCMs, such as quercetin also inactivated the SPI-1 T3SS and attenuated S. Typhimurium invasion. Our results demonstrate that specific plant metabolites from TCMs can directly interfere with key bacterial virulence pathways and reveals a previously unappreciated mechanism of action for anti-infective medicinal plants.
Pyrroloquinoline quinone [4,5-dihydro-4,5-dioxo-1H-pyrrolo[2,3-f]quinoline-2,7,9-tricarboxylic acid (PQQ)] is a bacterial vitamin that serves as a cofactor in numerous alcohol dehydrogenases. Its biosynthesis in Klebsiella pneumoniae is facilitated by six genes, pqqABCDEF, and proceeds by an unknown pathway. The protein encoded by pqqC catalyzes the final step of PQQ formation, which involves a ring closure and an overall eight-electron oxidation of 3a-(2-amino-2-carboxyethyl)-4,5-dioxo-4,5,6,7,8,9-hexahydroquinoline-7,9-dicarboxylic acid (AHQQ) in the absence of a redox-active metal or cofactor. A recent crystal structure has implicated numerous PQQ-PqqC interactions [Magnusson et al. (2004) Proc. Natl. Acad. Sci. U.S.A. 101, 7913-7918]. To investigate the mechanism of the PqqC reaction, the active site residue His84 has been mutated to H84A and H84N, and the kinetic and spectroscopic properties have been compared to each other and the wild-type enzyme using aerobic and anaerobic conditions. Both mutants form PQQ under aerobic conditions with rate constants of 0.09 min-1 and 0.056 min-1 relative to 0.34 min-1 for the wild-type enzyme. In addition to the initial E-AHQQ complex (532-536 nm) and the product E-PQQ complex (346-366 nm), a number of spectral intermediates are observed between 316 and 344 nm. The anaerobic reaction is particularly informative, showing that while mixing of H84N with AHQQ leads to a 344 nm intermediate, this is unable to proceed to a final 318 nm species; by contrast H84A forms the 344 nm species as a precursor to the 318 nm species. In the context of the proposed chemical mechanism for PqqC [Magnusson et al. (2004) Proc. Natl. Acad. Sci. U.S.A. 101, 7913-7918], we assign the 344 nm intermediate to a quinoid species and the 318 nm intermediate to an initial quinol species. The proposed role of H84 is as a proton donor to the oxyanion of the quinoid species such that subsequent C-H bond cleavage can occur to form a monoanionic quinol. In the absence of a proton donor (as occurs in H84N), the normal reaction path is precluded as this would require formation of an unstable, dianionic species. Unlike H84N, H84A appears to be small enough to allow entry of active site water, which is postulated to adopt the role of active site proton donor.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.