Discovery of Two Bacterial Nitric Oxide-Responsive Proteins and Their Roles in Bacterial Biofilm Regulation

Abstract

CONSPECTUS Bacterial biofilms form when bacteria adhere to a surface and produce an exopolysaccharide matrix (Costerton et al. Science 1999, 284, 1318; Davies et al. Science 1998, 280, 295; Flemming et al. Nat. Rev. Microbiol. 2010, 8, 623). Because biofilms are resistant to antibiotics, they are problematic in many aspects of human health and welfare, causing, for instance, persistent fouling of medical implants such as catheters and artificial joints (Brunetto et al. Chimia 2008, 62, 249). They are responsible for chronic infections in the lungs of cystic fibrosis patients and in open wounds, such as those associated with burns and diabetes. They are also a major contributor to hospital-acquired infections (Sievert et al. Infec. Control Hosp. Epidemiol. 2013, 34, 1; Tatterson et al. Front. Biosci. 2001, 6, D890). It has been hypothesized that effective methods of biofilm control will have widespread application (Landini et al. Appl. Microbiol. Biotechnol. 2010, 86, 813). A promising strategy is to target the mechanisms that drive biofilm dispersal, because dispersal results in biofilm removal and in the restoration of antibiotic sensitivity. First documented in Nitrosomonas europaea (Schmidt et al. J. Bacteriol. 2004, 186, 2781) and the cystic fibrosis-associated pathogen Pseudomonas aeruginosa (Barraud et al. J. Bacteriol. 2006, 188, 7344; J. Bacteriol. 2009, 191, 7333), regulation of biofilm formation by nanomolar levels of the diatomic gas nitric oxide (NO) has now been documented in numerous bacteria (Barraud et al. Microb. Biotechnol. 2009, 2, 370; McDougald et al. Nat. Rev. Microbiol. 2012, 10, 39; Arora et al. Biochemistry 2015, 54, 3717; Barraud et al. Curr. Pharm. Des. 2015, 21, 31). NO-mediated pathways are, therefore, promising candidates for biofilm regulation. Characterization of the NO sensors and NO-regulated signaling pathways should allow for rational manipulation of these pathways for therapeutic applications. Several laboratories, including our own, have shown that a class of NO sensors called H-NOX (heme-nitric oxide or oxygen binding domain) affects biofilm formation by regulating intracellular cyclic di-GMP concentrations and quorum sensing (Arora et al. Biochemistry 2015, 54, 3717; Plate et al. Trends Biochem. Sci. 2013, 38, 566; Nisbett et al. Biochemistry 2016, 55, 4873). Many bacteria that respond to NO do not encode an hnoX gene, however. My laboratory has now discovered an additional family of bacterial NO sensors, called NosP (nitric oxide sensing protein). Importantly, NosP domains are widely conserved in bacteria, especially Gram-negative bacteria, where they are encoded as fusions with or in close chromosomal proximity to histidine kinases or cyclic di-GMP synthesis or phosphodiesterase enzyme, consistent with signaling. In this Account, we briefly review NO and HNOX signaling in bacterial biofilms, describe our discovery of the NosP family, and provide support for its role in biofilm regulation in Pseudomonas aeruginosa, Vibrio cholerae, Legionella pneumophila, and Shewanella o...

Nitric Oxide Therapy for Diabetic Wound Healing

“…With additional protection afforded by an EPS, these communities of bacteria are highly resistant to traditional antibiotics . Though the exact mechanism of NO's antibiofilm action remains unclear, it is known that NO facilitates biofilm dispersal at low NO concentrations (10 −12 –10 −9 m ) and complete biofilm eradication at higher NO concentrations (10 −6 –10 −3 m ) . Large concentrations of NO from macrophage‐generated iNOS, therefore, act as a broad‐spectrum antimicrobial against both planktonic and biofilm‐based bacteria, reducing bacterial loads to a level at which normal wound healing processes are able to proceed …”

Nitric Oxide–Releasing Macromolecular Scaffolds for Antibacterial Applications

“…[16] Although the mechanisms or pathways of NO-regulated biofilm dispersal are still not clear, one popular hypothesis is that NO mediates the biofilm structure via cyclic-di-GMP (c-di-GMP), a signaling molecule involved in the production and maintenance of the biofilm EPS matrix. [15a, 17] Recent experimental evidence indicates the similarity of heme-NO/oxygen domains (H-NOX) and the genes coding for c-di-GMP cyclases or phosphodiesterases in bacteria, suggesting that the ligation of H-NOX with NO results in NO-mediated regulation of biofilms through a c-di-GMP signaling. [17] …”

“…[15a, 17] Recent experimental evidence indicates the similarity of heme-NO/oxygen domains (H-NOX) and the genes coding for c-di-GMP cyclases or phosphodiesterases in bacteria, suggesting that the ligation of H-NOX with NO results in NO-mediated regulation of biofilms through a c-di-GMP signaling. [17] …”

“…[15a,17] Recent experimental evidence indicates the similarity of hemeNO/oxygen domains (HNOX) and the genes coding for cdiGMP cyclases or phosphodiester ases in bacteria, suggesting that the ligation of HNOX with NO results in NOmediated regulation of biofilms through a cdiGMP signaling. [17] Barraud et al reported the benefit of low levels of exogenous NO on biofilm dispersal using low molecular weight NO donors. In an early study, NO was shown to disperse mature P. aeruginosa biofilms at ≈(0.025-0.50) × 10 −9 m over 24 h, ultimately initi ating detachment of bacteria without compromising the viability of planktonic bacteria.…”

Synthesis of the Ruthenium Nitrosyl Complex with Coordinated Ammonia and Pyridine at Room Temperature

“…Identification of nitric oxide (NO) as an endogenously produced diatomic free radical with various biochemical roles leads to the Nobel Prize in 1998 , . This radical is associated with a wide range of physiological roles, including platelet aggregation and adhesion, vasodilation,, wound repair, the immune response, and carcinogenesis , . Usage of various NO donors (nitroglycerin and etc.)…”