Biology subjects

Musayev, F. N.

Publications and source records attributed to Musayev, F. N..

4 recordsLinked to original sources

bioRxiv2026

Revealing interactions between glutathione peroxidase 4 and phosphoinositides

Glutathione peroxidase 4 (GPx4) is the primary enzyme reducing lipid hydroperoxides, preventing membrane oxidative damage and protecting against ferroptosis. GPx4 is known to engage with lipid headgroups through electrostatic interactions, positioning the substrate for reduction. This work reveals and characterizes binding of highly anionic phosphoinositides (PIP lipids) by GPx4. PIPs are vital lipids in human cells and are central to many signaling processes, particularly in cytosolic facing membranes. Lipid overlay assays confirm interactions between GPx4 and phosphorylated PIPs, comparable to known anionic lipid binders. Protein NMR describes the interaction between GPx4 and PIPs within micelles. The greatest resonance shifting occurs with trisphosphorylated PIP, suggesting that higher anionic charge leads to greater binding, a known driver of GPx4 substrate recognition. Preferred anionic interactions were also confirmed with titration and crystallographic structure analysis of inositol phosphate 4 (IP4). A headgroup-binding site on GPx4 is revealed to be proximal to the cationic membrane interaction site. In conjunction with molecular simulations, these results show that PIP lipid interactions allow full engagement of GPx4 with the membrane and positions the headgroup to allow the lipid tail to interact with the catalytic site. Understanding whether GPx4 preferentially interacts with PIPs will allow better understanding of the protective function of this essential enzyme and a mechanism that may protect essential lipid signaling pathways from oxidative damage. SignificanceThis study allows a deeper structural and mechanistic understanding of GPx4, the primary enzyme that reduces lipid hydroperoxides and prevents ferroptosis. Gaining an understanding of phosphoinositide binding to GPx4 reveals a mechanism for potential preservation of these important signaling molecules and for ferroptosis protection. Observation of a specific binding site for headgroup engagement reveals a plausible lipid interaction mode and functional mechanism of this important cytoprotective enzyme.

biophysics↗

bioRxiv2025

Broad PFAS binding with fatty acid binding protein 4 is enabled by variable binding modes.

Per- and polyfluoroalkyl substances (PFAS) are ubiquitous pollutants that bioaccumulate in wildlife and humans, yet the molecular basis of their protein interactions remains poorly understood. Here, we show that human adipocyte fatty acid-binding protein (FABP4) can bind a diverse array of PFAS, including next-generation replacements for legacy chemicals and longer-chain perfluorocarboxylic acids. Shorter-chain PFAS, although weaker binders, still displayed measurable affinities--surpassing those of their nonfluorinated analogs. We determined crystal structures of FABP4 bound to perfluorooctanoic acid (PFOA), perfluorodecanoic acid (PFDA), and perfluorohexadecanoic acid (PFHxDA), revealing three distinct binding modes. Notably, PFOA binds in two separate sites, and two distinct conformations define single-ligand binding of PFDA and PFHxDA. These arrangements enhance hydrophobic interactions within the binding cavity and likely explain the low micromolar dissociation constants observed in fluorescence competition assays. Our findings underscore the critical roles of chain length, headgroup functionality, and protein conformation in PFAS-FABP4 interactions. Given the emerging implications of the role of FABP4 in endocrine function, even subtle PFAS-induced perturbations could affect metabolic regulation and disease risk. Overall, this work highlights the value of direct structural and biochemical insights into PFAS-FABP4 interactions and paves the way for future research on PFAS transport and toxicological outcomes.

biophysics↗

bioRxiv2024

Full-length structure and heme binding in the transcriptional regulator HcpR

HcpR is a CRP-family transcriptional regulator found in many Gram-negative anaerobic bacteria. In the perio-pathogen Porphyromonas gingivalis, HcpR is crucial for the response to reactive nitrogen species such as nitric oxide (NO). Binding of NO to the heme group of HcpR leads to transcription of the redox enzyme Hcp. However, the molecular mechanisms of heme binding to HcpR remain unknown. In this study we present the 2.3[A] structure of the P. gingivalis HcpR. Interdomain interactions present in the structure help to form a hydrophobic pocket in the N-terminal sensing domain. A comparison analysis with other CRP-family members reveals that the molecular mechanisms of HcpR-mediated regulation may be distinct from other family members. Using docking studies, we identify a putative heme binding site in the sensing domain. In vitro complementation and mutagenesis studies verify Met68 as an important residue in activation of HcpR. Finally, heme binding studies with purified forms of recombinant HcpR support Met68 and His149 residues as important for proper heme coordination in HcpR.

microbiology↗

bioRxiv2020

The Cryo-EM structure of AAV2 Rep68 in complex with ssDNA reveals a malleable AAA+ machine that can switch between oligomeric states.

The adeno-associated virus (AAV) Rep proteins use a unique AAA+ domain to catalyze DNA replication, transcription regulation, and genome packaging. Also, they mediate site-specific integration during a latent phase. To understand the mechanisms underlying AAV Rep function, we investigated the cryo-EM and X-ray structures of Rep68-ssDNA complexes. Surprisingly, Rep68 generates hybrid ring structures where the Origin-Binding-Domain (OBD) forms octameric rings while the helicase domain (HD) forms heptamers. Moreover, the binding to ATP{gamma}S promotes a large conformational change in the entire AAA+ domain that leads the HD to form both heptamer and hexamers. The HD oligomerization is driven by an interdomain linker region that acts as a latch to catch the neighboring HD subunit and is flexible enough to permit the formation of different stoichiometric ring structures. Overall, our studies show the structural basis of AAV Reps structural flexibility required to fulfill its multifunctional role during the AAV life cycle.

biochemistry↗