Biology subjects

Kroll, T.

Publications and source records attributed to Kroll, T..

7 recordsLinked to original sources

bioRxiv2025

Mapping the Exit Route of Hydrogen Peroxide From the Manganese Superoxide Dismutase (MnSOD) Active Site.

Human mitochondrial manganese superoxide dismutase (MnSOD) converts superoxide (O [bullet]-) into hydrogen peroxide (H O ) and molecular oxygen (O ), serving as a key defense against oxidative damage. Despite extensive studies, the full structural characterization of H2O2-binding sites in MnSOD remains largely unexplored. Previous H2O2-soaked MnSOD structures have identified two distinct H2O2-binding sites: one directly ligated to the catalytic Mn (LIG position) and another at the active site gateway (PEO position) between second-shell residues Tyr34 and His30. In this study, a kinetically impaired Gln143Asn MnSOD variant is used to trap and explore additional H2O2-binding sites beyond the second-shell solvent gate. In the wild-type enzyme, Gln143 mediates proton transfers with the Mn-bound solvent (WAT1) to drive redox cycling of the metal, necessary for effective O [bullet]- dismutation. Substitution with Asn stalls catalysis because the increased distance from WAT1 disrupts critical proton-coupled electron transfer (PCET) events, and the redox cycling of the active site metal is impaired. This, in turn, stalls the electrostatic cycling of positive charge on the enzyme surface and enhances the likelihood of trapping transient H2O2-bound states in this variant. Results reveal several H2O2 molecules leading up to the active site, in addition to the canonical LIG and PEO positions. SynopsisA high-resolution X-ray structure of a Gln143Asn variant of manganese superoxide dismutase reveals multiple hydrogen peroxide binding sites beyond the canonical LIG and PEO binding positions within the active site. These findings expand the known landscape of product peroxide-bound states in MnSOD.

biochemistry↗

bioRxiv2024

The role of Tyr34 in proton-coupled electron transfer of human manganese superoxide dismutase

Human manganese superoxide dismutase (MnSOD) plays a crucial role in controlling levels of reactive oxygen species (ROS) by converting superoxide (O2*-) to molecular oxygen (O2) and hydrogen peroxide (H2O2) with proton-coupled electron transfers (PCETs). The reactivity of human MnSOD is determined by the state of a key catalytic residue, Tyr34, that becomes post-translationally inactivated by nitration in various diseases associated with mitochondrial dysfunction. We previously reported that Tyr34 has an unusual pKa due to its proximity to the Mn metal and undergoes cyclic deprotonation and protonation events to promote the electron transfers of MnSOD. To shed light on the role of Tyr34 MnSOD catalysis, we performed neutron diffraction, X-ray spectroscopy, and quantum chemistry calculations of Tyr34Phe MnSOD in various enzymatic states. The data identifies the contributions of Tyr34 in MnSOD activity that support mitochondrial function and presents a thorough characterization of how a single tyrosine modulates PCET catalysis.

biochemistry↗

bioRxiv2024

Revealing the atomic and electronic mechanism of human manganese superoxide dismutase product inhibition

Human manganese superoxide dismutase (MnSOD) is a crucial oxidoreductase that maintains the vitality of mitochondria by converting O2[bullet]- to O2 and H2O2 with proton-coupled electron transfers (PCETs). Since changes in mitochondrial H2O2 concentrations are capable of stimulating apoptotic signaling pathways, human MnSOD has evolutionarily gained the ability to be highly inhibited by its own product, H2O2. A separate set of PCETs is thought to regulate product inhibition, though mechanisms of PCETs are typically unknown due to difficulties in detecting the protonation states of specific residues that coincide with the electronic state of the redox center. To shed light on the underlying mechanism, we combined neutron diffraction and X-ray absorption spectroscopy of the product-bound, trivalent, and divalent states to reveal the all-atom structures and electronic configuration of the metal. The data identifies the product-inhibited complex for the first time and a PCET mechanism of inhibition is constructed.

biochemistry↗

bioRxiv2023

Orphan nuclear receptor NR2E3 and its small-molecule agonist induce cancer cell apoptosis through regulating p53, IFNα and MYC pathways

Orphan nuclear receptor NR2E3 activates p53 and induces cancer cell apoptosis. Further studies on p53-dependent and -independent functions of wild-type and mutated NR2E3 are needed. Herein, we showed that NR2E3 enhanced p53-DNA interactions in diverse cancer cells and up-regulated p53 and IFN pathways while down-regulating MYC pathway in cervical cancer cells. Studies of "All of Us" and TCGA databases showed NR2E3 nonsynonymous mutations associating with four cancers. We stratified NR2E3 SNVs for their cancer implications with the p53 reporter. A cancer-associated NR2E3R97Hmutation not only lost the wild-types tumor-suppressing functions but also prohibited the wild-type from enhancing p53 acetylation. These observations implicated the potential for pharmaceutically activating NR2E3 to suppress cancer. Indeed, NR2E3s small-molecule agonist 11a repressed 2-D and 3-D cultures of primary cells and cell lines of cervical cancer, in which screening FDA-approved anti-cancer drugs identified HDAC-1/2 inhibitor Romidepsin operating synergistically with 11a. The underlying molecular mechanisms included 11as down-regulating the transcription of Multidrug Resistance Protein ABCB1 that Romidepsin up-regulated. Transcriptomics studies revealed three synergy modes: (1) "sum-up" mode that the p53 pathway activated individually by 11a and Romidepsin got stronger by the combo; (2) "antagonism" mode that Romidepsin counteracted the activation of the Kras pathway by 11a; and (3) "de novo" mode that the combo instead of each individual drug repressed the MYC pathway. Conclusively, our experiments provide new data supporting tumor-suppressor like functions for wild-type NR2E3, reveal roles of mutated NR2E3 in cancer, and address values of NR2E3s agonist 11a in cancer therapy alone and combined.

cancer biology↗

bioRxiv2023

A genetic variation in the adenosine A2A receptor gene contributes to variability in oscillatory alpha power in wake and sleep EEG and A1 adenosine receptor availability in the human brain

The EEG alpha rhythm (8-13 Hz) is one of the most salient human brain activity rhythms. Spectral power in the alpha range in wakefulness and sleep varies among individuals based on genetical predisposition, yet knowledge about the underlying genes is scarce. The EEG alpha oscillations are related to cerebral energy metabolism and modulated by the level of attention and vigilance. The neuromodulator adenosine is directly linked to energy metabolism as product of adenosine tri-phosphate (ATP) breakdown and acts as a sleep promoting molecule by activitating A1 and A2A adenosine receptors. We quantified EEG oscillatory alpha power in wakefulness and sleep, as well as A1 adenosine receptor availability by positron emission tomography with 18F-CPFPX, in a large sample of healthy volunteers carrying different alleles of gene variant rs5751876 of ADORA2A encoding A2A adenosine receptors. Oscillatory alpha power was higher in homozygous C-allele carriers (n = 27, 11 females) compared to heterozygous and homozygous carriers of the T-allele (n(C/T) = 23, n(T/T) = 5, 13 females) (F(18,37) = 2.35, p = 0.014, Wilks {Lambda} = 0.467). Across considered brain regions an effect of ADORA2A genotype on A1 adenosine receptor binding potential was found (F(18,40) = 2.62, p = 0.006, Wilks {Lambda} = 0.459) and after correction for multiple testing this effect was shown to be significant for circumscribed occipital region of calcarine fissures. A correlation between individual differences in oscillatory alpha power and adenosine receptor availability was found for the subgroup of female participants only. In conclusion: a genetic variation in the adenosinergic system affects individual alpha power, although a direct modulatory effect via the A1AR has been demonstrated for females only.

neuroscience↗

bioRxiv2021

Isolation, omics characterization and organotypic culture of alveolar type II pulmonary epithelial cells

The alveolar type II (AT2) epithelial cell fraction includes the stem cells of the pulmonary alveoli, functioning in lung homeostasis and post-injury repair. AT2 cells have been characterized primarily in situ, in transgenic mouse models. We report a new methodology for their isolation, their "omics" characterization and stroma-cell-free organotypic culture. Our multi-omics analysis identified high expression of genes involved in oxidative phosphorylation and of AP-1 components, as well as new phosphorylation sites in AT2 biomarkers. Furthermore, we show that supplementation with KGF, FGF10 & HGF suffices for the in vitro proliferation of AT2 cells and formation of alveolar organoids, suggesting that AT2-based organotypic development depends on ligands of the c-Met and FGFR2 receptors. The reported methodology and in-depth molecular characterization provide new tools for the in vitro and in vivo functional analysis of pulmonary cells and of mouse models of lung disease.

cell biology↗

bioRxiv2017

Zinc storage granules in the Malpighian tubules of Drosophila melanogaster

Membrane transporters and sequestration mechanisms concentrate metal ions differentially into discrete subcellular microenvironments for usage in protein cofactors, signaling, storage, or excretion. Here we identify zinc storage granules as the insects major zinc reservoir in primary Malpighian tubule epithelial cells of Drosophila melanogaster. The concerted action of Adaptor Protein-3, Rab32, HOPS and BLOC complexes as well as of the white-scarlet (ABCG2-like) and ZnT35C transporters is required for zinc storage granule biogenesis. Due to similar lysosome related organelle defects, patients with Hermansky-Pudlak syndrome may lack zinc granules in beta pancreatic cells, intestinal paneth cells and presynaptic vesicles of hippocampal mossy fibers.

genetics↗