Detecting and Understanding Nitric Oxide Formation during Nitrogen Cycling in Microbial Biofilms (2009)

Nitric Oxide (NO) is a remarkable molecule and, even though not obvious for us, it is ubiquitously present in low concentrations.It is essential for biogeochemical nitrogen cycling in sediments and waters, an important component of the atmospheric chemistry, and it regulates many functions in microorganisms and in multicellular organisms including humans.The discovery that NO is a central signaling molecule for the regulation of blood pressure sparked its fame, despite of being a simple gaseous molecule, resulting in its selection as the molecule of the year in 1992, followed by the award of the Nobel Prize in Physiology and Medicine in 1998 for the main investigators involved (R.F.Furchgott; F. Murad, and L.J. Ignarro).However, it was already recognized by T. Yamanaka in 1956 that NO was an active biomolecule.He showed NO formation by cell-free extracts that contained bacterial cytochromes and hypothesized that it is an intermediate in denitrification.This hypothesis was given wide acceptance when bacterial NO reductase of denitrifiers was isolated at the end of 1980ies.Somewhat later (1993) the signaling role of NO in bacteria was established by showing its involvement in regulating the oxidative stress response mediated by the SoxRS system, which was followed by many other examples.This thesis started with the ambitious goal of establishing a signaling role for NO in complex microbial communities organized in biofilms.Before assessing the signaling role of NO, however, I had to realize that even reliable detection of NO in these environments at the relevant scale was not possible.Moreover, the manifold reactions of the nitrogen cycle that can possibly produce NO cause a considerable uncertainty about the processes that are responsible for NO formation.Thus, I learned a lesson every researcher receives: "Go to the lab and eat humble pie, but keep your big ideas in mind."Literally, this meant the development of an NO microsensor that is suitable for measurements in biofilms and sediments, and its application to complex microbial communities in dental and nitrifying/denitrifying biofilms.Application of the NO microsensor, together with other microsensors, resulted in knowledge of the microenvironment and the activity of nitrogen cycling pathways, and allowed the rather firm assignment of accompanied NO formation to specific pathways.But what about the 'big ideas'?Following the transient formation of NO in the nitrifying/denitrifying biofilm suggested that NO formation overshoots during perturbation of an active nitrogen cycling pathway.This overshoot is counteracted immediately, possibly on the enzyme level within this complex community.This is far from proving a signaling function but it shows a high degree of regulation of free NO concentrations and highlights the importance of NO for the functioning of the whole community.NO formation by the dental microbial community led us to the exciting hypothesis that a human-associated biofilm community may interact with the host via NO, because NO is known to be involved in shaping the immune response in the gum.The definitive establishment of NO signaling within and out of complex microbial communities organized in biofilms clearly awaits a concerted effort of microbial ecologists and molecular biologists in the future.This is certainly a task which could fill a researcher's life.