top of page
Publications
A collection of research and review articles published by members of the action
The [2Fe-2S] cluster of mitochondrial outer membrane protein mitoNEET has an O2-regulated nitric oxide access tunnel
The mitochondrial outer membrane iron–sulphur ([Fe-S]) protein mitoNEET has been extensively studied as a target of the anti-inflammatory and type-2 diabetes drug pioglitazone and as a protein affecting mitochondrial respiratory rate. Despite these extensive past studies, its molecular function has yet to be discovered. Here, we applied an interdisciplinary approach and discovered an explicit nitric oxide (NO) access site to the mitoNEET [2Fe-2S] cluster. We found that O2 and pioglitazone block NO access to the cluster, suggesting a molecular function for the mitoNEET [2Fe-2S] cluster in mitochondrial signal transduction. Our discovery hints at a new pathway via which mitochondria can sense hypoxia through O2 protection of the mitoNEET [2Fe-2S] cluster, a new paradigm in understanding the importance of [Fe-S] clusters for gasotransmitter signal transduction in eukaryotes.
A Versatile Virus-Mimetic Engineering Approach for Concurrent Protein Nanocage Surface-Functionalization and Cargo Encapsulation
Naturally occurring protein nanocages like ferritin are self-assembled from multiple subunits. Because of their unique cage-like structure and biocompatibility, there is a growing interest in their biomedical use. A multipurpose and straightforward engineering approach does not exist for using nanocages to make drug-delivery systems by encapsulating hydrophilic or hydrophobic drugs and developing vaccines by surface functionalization with a protein like an antigen. Here, a versatile engineering approach is described by mimicking the HIV-1 Gap polyprotein precursor. Various PREcursors of nanoCages (PREC) are designed and created by linking two ferritin subunits via a flexible linker peptide containing a protease cleavage site. These precursors can have additional proteins at their N-terminus, and their protease cleavage generates ferritin-like nanocages named protease-induced nanocages (PINCs). It is demonstrated that PINC formation allows concurrent surface decoration with a protein and hydrophilic or hydrophobic drug encapsulation up to fourfold more than the amount achieved using other methods. The PINCs/Drug complex is stable and efficiently kills cancer cells. This work provides insight into the precursors’ design rules and the mechanism of PINCs formation. The engineering approach and mechanistic insight described here will facilitate nanocages’ applications in drug delivery or as a platform for making multifunctional therapeutics like mosaic vaccines.
Biochemical and cellular characterization of the CISD3 protein: Molecular bases of cluster release and destabilizing effects of nitric oxide
The NEET proteins, an important family of iron-sulfur (Fe-S) proteins, have generated a strong interest due to their involvement in diverse diseases such as cancer, diabetes, and neurodegenerative disorders. Among the human NEET proteins, CISD3 has been the least studied, and its functional role is still largely unknown. We have investigated the biochemical features of CISD3 at the atomic and in cellulo levels upon challenge with different stress conditions i.e., iron deficiency, exposure to hydrogen peroxide, and nitric oxide. The redox and cellular stability properties of the protein agree on a predominance of reduced form of CISD3 in the cells. Upon the addition of iron chelators, CISD3 loses its Fe-S clusters and becomes unstructured, and its cellular level drastically decreases. Chemical shift perturbation measurements suggest that, upon cluster oxidation, the protein undergoes a conformational change at the C-terminal CDGSH domain, which determines the instability of the oxidized state. This redox-associated conformational change may be the source of cooperative electron transfer via the two [Fe2S2] clusters in CISD3, which displays a single sharp voltammetric signal at −31 mV versus SHE. Oxidized CISD3 is particularly sensitive to the presence of hydrogen peroxide in vitro, whereas only the reduced form is able to bind nitric oxide. Paramagnetic NMR provides clear evidence that, upon NO binding, the cluster is disassembled but iron ions are still bound to the protein. Accordingly, in cellulo CISD3 is unaffected by oxidative stress induced by hydrogen peroxide but it becomes highly unstable in response to nitric oxide treatment.
Chelation of Mitochondrial Iron as an Antiparasitic Strategy
Iron, as an essential micronutrient, plays a crucial role in host–pathogen interactions. In order to limit the growth of the pathogen, a common strategy of innate immunity includes withdrawing available iron to interfere with the cellular processes of the microorganism. Against that, unicellular parasites have developed powerful strategies to scavenge iron, despite the effort of the host. Iron-sequestering compounds, such as the approved and potent chelator deferoxamine (DFO), are considered a viable option for therapeutic intervention. Since iron is heavily utilized in the mitochondrion, targeting iron chelators in this organelle could constitute an effective therapeutic strategy. This work presents mitochondrially targeted DFO, mitoDFO, as a candidate against a range of unicellular parasites with promising in vitro efficiency. Intracellular Leishmania infection can be cleared by this compound, and experimentation with Trypanosoma brucei 427 elucidates its possible mode of action. The compound not only affects iron homeostasis but also alters the physiochemical properties of the inner mitochondrial membrane, resulting in a loss of function. Furthermore, investigating the virulence factors of pathogenic yeasts confirms that mitoDFO is a viable candidate for therapeutic intervention against a wide spectrum of microbe-associated diseases.
Ancient complexes of iron and sulfur modulate oncogenes and oncometabolism
Inorganic complexes of iron and sulfur, that is, iron-sulfur [FeS] clusters, have played a fundamental role in life on Earth since the prebiotic period. These clusters were involved in elementary reactions leading to the emergence of life and, since then, gained function in processes, such as respiration, replication, transcription, and the immune response. We discuss how three [FeS] proteins involved in the innate immune response play a role in oncogene expression/function and oncometabolism. Our analysis highlights the importance of future research into understanding the [FeS] clusters' roles in cancer progression and proliferation. The outcomes of these studies will help identify new targets and develop new anticancer therapeutics.
VITAS, a sensitive in vivo selection assay to discover enzymes producing antiviral natural products
To discover new broad-spectrum antiviral nucleotide analogues from natural resources or through protein engineering, we have developed a sensitive in vivo selection assay named Viral polymerase-Inhibition Toxin-Associated Selection (VITAS). We show that the assay works with enzymes from three Kingdoms of life.
Radical-SAM dependent nucleotide dehydratase (SAND), rectification of the names of an ancient iron-sulfur enzyme using NC-IUBMB recommendations
In 1789, the influential French chemist Antoine-Laurent Lavoisier described his view of science and its langague in his book Traité élémentaire de chimie. According to the Robert Kerr’s translation it states (Lavoisier, 1790): “As ideas are preserved and communicated by means of words, it necessarily follows that we cannot improve the language of any science without at the same time improving the science itself; neither can we, on the other hand, improve a science without improving the language or nomenclature which belongs to it.” This view reminds us of Confucius’s earlier doctrine, the rectification of names (Steinkraus, 1980; Lau, 2000). Confucius believed that rectification of names is imperative. He explained (Steinkraus, 1980; Lau, 2000): “If language is incorrect, then what is said does not concord with what was meant, what is to be done cannot be affected. If what is to be done cannot be affected, then rites and music will not flourish. If rites and music do not flourish, then mutilations and lesser punishments will go astray. And if mutilations and lesser punishments go astray, then the people have nowhere to put hand or foot. Therefore the gentleman uses only such language as is proper for speech, and only speaks of what it would be proper to carry into effect. The gentleman in what he says leaves nothing to mere chance.” Inspired by these views, we make the analogy that the progress of science and the language used to describe it are two entangled electrons. This entanglement highlights the importance of introducing systemic names for enzymes using EC classification and the ever-growing problem of protein names (McDonald and Tipton, 2021). Here, we tackle one specific case of iron-sulfur ([FeS]) enzymes. We show that the language used to describe a conserved [FeS] enzyme of the innate immune system, i.e., viperin or RSAD2, is now inadequate and disentangled from its science. We discuss that the enzyme has cellular functions beyond its antiviral activity and that eukaryotic and prokaryotic enzymes catalyse the same chemical reactions. To prevent bias towards antiviral activity while studying various biochemical activities of the enzyme and using scientifically incorrect terms like “prokaryotic viperins,” we rectify the language describing the enzyme. Based on NC-IUBMB recommendations, we introduce the nomenclature S-adenosylmethionine (SAM) dependent Nucleotide Dehydratase (SAND).
bottom of page