2-Nitropropane dioxygenase (EC 1.13.11.32) catalyzes the oxidation of nitroalkanes into their corresponding carbonyl compounds and nitrite. In this study, the ncd-2 gene encoding for the enzyme in Neurospora crassa was cloned, expressed in Escherichia coli, and the resulting enzyme was purified. Size exclusion chromatography, heat denaturation, and mass spectroscopic analyses showed that 2-nitropropane dioxygenase is a homodimer of 80 kDa, containing a mole of non-covalently bound FMN per mole of subunit, and is devoid of iron. With neutral nitroalkanes and anionic nitronates other than propyl-1-and propyl-2-nitronate, for which a nonenzymatic free radical reaction involving superoxide was established using superoxide dismutase, substrate oxidation occurs within the enzyme active site. The enzyme was more specific for nitronates than nitroalkanes, as suggested by the second order rate constant k cat /K m determined with 2-nitropropane and primary nitroalkanes with alkyl chain lengths between 2 and 6 carbons. The steady state kinetic mechanism with 2-nitropropane, nitroethane, nitrobutane, and nitrohexane, in either the neutral or anionic form, was determined to be sequential, consistent with oxygen reacting with a reduced form of enzyme before release of the carbonyl product. Enzyme-monitored turnover with ethyl nitronate as substrate indicated that the catalytically relevant reduced form of enzyme is an anionic flavin semiquinone, whose formation requires the substrate, but not molecular oxygen, as suggested by anaerobic substrate reduction with nitroethane or ethyl nitronate. Substrate deuterium kinetic isotope effects with 1,2-[ 2 H 4 ]nitroethane and 1,1,2-[ 2 H 3 ]ethyl nitronate at pH 8 yielded normal and inverse effects on the k cat /K m value, respectively, and were negligible on the k cat value. The k cat /K m and k cat pH profiles with anionic nitronates showed the requirement of an acid, whereas those for neutral nitroalkanes were consistent with the involvement of both an acid and a base in catalysis. The kinetic data reported herein are consistent with an oxidasestyle catalytic mechanism for 2-nitropropane dioxygenase, in which the flavin-mediated oxidation of the anionic nitronates or neutral nitroalkanes and the subsequent oxidation of the enzyme-bound flavin occur in two independent steps.2-Nitropropane dioxygenase (EC 1.13.11.32) from Neurospora crassa is a flavin-dependent enzyme, which catalyzes the oxidative denitrification of nitroalkanes to their corresponding carbonyl compounds and nitrite. The enzyme was first characterized by Little (1) in 1951 as an oxidase, but was later classified as a dioxygenase based on the observation that the oxygen atom of the organic product formed upon oxidation of propyl-2-nitronate originates from molecular oxygen and not from water (2). To date, 2-nitropropane dioxygenase has been isolated from N. crassa (1) and Hansenula mrakii (3). Whereas the two enzymes have similar molecular weights of about 40,000, they differ in their prosthetic group content in that...

Background Connected medical devices and electronic health records have added important functionality to patient care, but have also introduced a range of cybersecurity concerns. When a healthcare organization suffers from a cybersecurity incident, its incident response strategies are critical to the success of its recovery. Objective In this article, we identify gaps in research concerning cybersecurity response plans in healthcare. Through a systematic literature review, we develop aggregated strategies that professionals can use to construct better response strategies in their organizations. Methods We reviewed journal articles on cyber incident response plans in healthcare published in PubMed and Web of Science. We sought to collect articles on the intersection of cybersecurity and healthcare that focused on incident response strategies. Results We identified and reviewed 13 articles for cybersecurity response recommendations. We then extracted information such as research methods, findings, and implications. Finally, we synthesized the recommendations into a framework of eight aggregated response strategies (EARS) that fall under managerial and technological categories. Conclusions We conducted a systematic review of the literature on cybersecurity response plans in healthcare and developed a novel framework for response strategies that could be deployed by healthcare organizations. More work is needed to evaluate incident response strategies in healthcare.

Psychological distress affects a significant proportion of caregivers of patients living longer with HGG. The presence of caregiver strain and low patient functional well-being are the most important predictors of psychological distress among this cohort of caregivers.

The process of triage in the palliative care setting is complex but can be conceptualised using a limited number of domains. Further research is required to establish the relative value clinicians attribute to these domains and thus inform the development of an acceptable and useful evidence-based palliative care triage tool.