Significance Heme protein sensors interact with various gaseous molecules, such as CO, NO, or O 2 , and play a crucial role in transcriptional and regulatory events. In general, the sensory domains of heme proteins control signal transduction domains such as histidine kinases, phosphodiesterases, DNA-binding domains, guanylate cyclases, diguanylate cyclase, and aerotaxis transducers. Here we report globin-coupled heme containing adenylate cyclase from Leishmania major (HemAC-Lm), which regulates O 2 -dependent cAMP synthesis. Oxygen binding at heme iron of HemAC-Lm presumably triggers a conformational change in the sensor domain that sequentially stimulates the catalytic activity of the adenylate cyclase domain, resulting in the synthesis of the second messenger cAMP. This O 2 -dependent cAMP signaling is likely to function in cellular adaptability during hypoxia.

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Background: NAD(P)H cytochrome b 5 oxidoreductase (Ncb5or) participates in a variety of metabolic conversions. Results: A new Ncb5or acts as a redox partner of ⌬12 fatty acid desaturase. Conclusion: Ncb5or null mutant suffers from impaired linoleate synthesis leading to oxidative stress and cell death. Significance: Our results have exposed a novel fatty acid synthesis pathway in L. major that differs from the mammalian system.

Previous optical and electron paramagnetic resonance (EPR) spectroscopic studies of the newly discovered peroxynitrite scavenging pseudoperoxidase from Leishmania major (LmPP) suggested that ferric LmPP contained a six-coordinate low-spin (6cLS) heme with a thiolate ligand, presumably a cysteine, bound to its heme iron. To identify the axial ligands of LmPP, we exploit a systematic mutational analysis of potential heme ligands. On the basis of UV-visible and EPR spectroscopy, we report that the substitution of the proximal His206 with alanine in LmPP alters the 6cLS to a five-coordinate high spin (5cHS) form at pH 4.0 that has a spectrum characteristic of a Cys-ligated 5cHS derivative. The electronic absorption and EPR analysis of all alanine-substituted Cys and Met single mutants establish that when Cys107 is replaced with alanine, a new species appears that has a spectrum characteristic of a histidine-ligated 5cHS derivative at pH 4.0. Together, these results suggest that His206 and Cys107 act as the proximal and distal axial ligands in ferric LmPP, respectively. However, the electronic properties of reduced wild-type LmPP are similar to those of known 5cHS His-ligated heme proteins at pH 8.8, indicating that the thiolate bond was broken upon reduction. Furthermore, the wild-type protein was only partially reduced at pH 4.0, but the E105L mutant was completely reduced to form a 5cHS ferrous heme. These results imply that the presence of an acidic residue near the distal site may prevent reduction of the heme iron at acidic pH.

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Severe COVID-19 frequently features a systemic deluge of cytokines. Circulating cytokines that can stratify risks are useful for more effective triage and management. Here, we ran a machine-learning algorithm on a dataset of 36 plasma cytokines in a cohort of severe COVID-19 to identify cytokine/s useful for describing the dynamic clinical state in multiple regression analysis. We performed RNA-sequencing of circulating blood cells collected at different time-points. From a Bayesian Information Criterion analysis, a combination of interleukin-8 (IL-8), Eotaxin, and Interferon-γ (IFNγ) was found to be significantly linked to blood oxygenation over seven days. Individually testing the cytokines in receiver operator characteristics analyses identified IL-8 as a strong stratifier for clinical outcomes. Circulating IL-8 dynamics paralleled disease course. We also revealed key transitions in immune transcriptome in patients stratified for circulating IL-8 at three time-points. The study identifies plasma IL-8 as a key pathogenic cytokine linking systemic hyper-inflammation to the clinical outcomes in COVID-19.

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