Biology subjects

Fahmy, K.

Publications and source records attributed to Fahmy, K..

7 recordsLinked to original sources

bioRxiv2026

Membrane mimetics control internal hydration and catalytic activity of the copper ATPase LpCopA from Legionella pneumophila

Abstract textMembrane protein function depends critically on the surrounding lipid bilayer, which supports specific lipid-protein interactions and imposes constraints through its ensemble properties. Ion-transporting ATPases undergo large conformational changes that perturb these interactions, and the high chemical potential of water can further drive conformational transitions through changes in internal hydration. Here, we investigated the relationship between membrane environment, internal hydration, and catalytic activity in the P1B-type copper ATPase LpCopA from Legionella pneumophila. BADAN-labeled LpCopA mutants revealed distinct basal hydration states near the conserved canonical binding site (CBS) cysteines C382 and C384, with C382 residing in a more hydrated environment than C384. Using the osmolyte PEG-1500, the spectral response of BADAN indicated an average internal hydration volume of about 800 [A]3 for LpCopA in E. coli lipid-doped mixed micelles (MMs), corresponding to approximately 25 water molecules. In nanodiscs (NDs), less than half of this volume responded to osmotic pressure, consistent with compaction of the transmembrane helical bundle under membrane lateral pressure. Remarkably, ATPase activity measured in different lipid reconstitution systems scaled with the extent of internal hydration, with diisobutylene/maleic acid lipid particles (DIBMALPs) imposing the tightest transmembrane compaction and lowest basal activity. These data indicate that the packing density and internal hydration of the transmembrane domain of LpCopA are strongly modulated by the membrane environment. The energetic estimates further support a model in which hydration-dependent expansion of the transmembrane domain against membrane lateral pressure contributes to the free-energy barrier of ATP hydrolysis.

biophysics↗

bioRxiv2025

Metabolic heat flow from the minimal cell JCVI-Syn3B reveals the lipidome-dependence of growth and metabolism

The cell membrane facilitates interactions with the environment and serves as an organizational platform for coordinating cellular processes, with lipids playing a central role in determining membrane property and function. Yet, how lipidome composition influences cellular fitness remains poorly defined. Recent approaches to chemically tune and minimize the lipidomes of genomically minimized bacterial organisms such as JCVI-Syn3A/B offer a streamlined system to explore why cells need such diverse lipid chemistries. In this study, we use isothermal microcalorimetry to assess how changes in lipid composition affect heat dissipated by JCVI-Syn3B cells, a parameter reflecting both growth and metabolic efficiency. By transposing the Monod equation into a calorimetric equation and extending it to the full life times of batch cultures, we introduce a new approach to quantify the metabolic efficiency of JCVI-Syn3B. Remarkably, our results demonstrate that tuning lipidome composition results in considerable variations of energy dissipation at the expense of biomass production. As a consequence, the volume of these minimal cells becomes inversely coupled to the lipidome-dependent entropic cost of cell division. The corresponding change in heat flow per cell mass gives rise to a complex but systematic dependence of growth rates on lipid composition. Interestingly, the maximal rate correlates with maximal lipid diversity, suggesting that the ability to tune local cell membrane charge and curvature through lipid structural diversity is crucial for divisome function. Our observations highlight the critical role of lipidome composition in cell metabolism and growth, and provide a new tool for interrogating the relationship between membrane composition and cell fitness.

biophysics↗

bioRxiv2025

X-ray Spectroscopy Meets Native Mass Spectrometry: Probing Gas-phase Protein Complexes

Gas-phase activation and dissociation studies of biomolecules, proteins and their non-covalent complexes using X-rays hold great promise for revealing new insights into the structure and function of biological samples. This is due to the unique properties of X-ray molecular interactions, such as site-specific and rapid ionization. In this perspective, we report and discuss the promise of first proof-of-principle studies of X-ray-induced dissociation of native biological samples ranging from small 17 kDa monomeric proteins up to large 808 kDa non-covalent protein assemblies conducted at a synchrotron (PETRA III) and a free-electron laser (FLASH2). A commercially available quadrupole time-of-flight mass spectrometer (Q-ToF2, Micromass/Waters), modified for high-mass analysis by MS Vision, was further adapted for integration with the open ports at the corresponding beamlines. The protein complexes were transferred natively into the gas phase via nano-electrospray ionization and subsequently probed by extreme ultraviolet (FLASH2) or soft X-ray (PETRA III) radiation, in either their folded state or following collision-induced activation in the gas phase. Depending on the size of the biomolecule and the activation method, protein fragmentation, dissociation, or enhanced ionization were observed. Additionally, an extension of the setup by ion mobility is described, which can serve as a powerful tool for structural separation of biomolecules prior to X-ray probing. The first experimental results are discussed in the broader context of current and upcoming X-ray sources, highlighting their potential for advancing structural biology in the future.

biochemistry↗

bioRxiv2024

Chemically defined lipid diets reveal the versatility of lipidome remodeling in genomically minimal cells

All cells are encapsulated in a lipid membrane that provides a responsive interface between life and its environment. Although simple membranes can be built from a single type of lipid, cellular membranes contain 10s to 100s of unique lipid species. Deciphering the significance of lipidome complexity is a central challenge in understanding the design principles of living membranes. While functions of individual lipids have been extensively studied, understanding how lipidomes collectively contribute to membrane function and cell phenotypes is experimentally challenging in most organisms. To address this challenge, we turned to the simple pathogenic organism Mycoplasma mycoides and its genomically derived "Minimal Cell" JCVI-syn3B, to establish a living minimal membrane model system in which lipidome complexity can be experimentally manipulated. By complexing lipids with cyclodextrins, we introduce a chemically defined approach to deliver lipid diets with different chemistries to cells, resulting in cellular lipidomes with as few as seven to nearly 30 lipids species. We explored how lipidome size and composition influences cell growth, osmotic sensitivity, and membrane adaptability to changes in growth temperature. Our findings indicate that lipidome composition dictates membrane adaptation to temperature change. Moreover, we show that lipidome diversity enhances cellular robustness to hypoosmotic shock. We further show that impaired acyl chain remodeling in the minimal cell is associated with impaired membrane temperature adaptation. Finally, we demonstrate as a proof of principle, how cells with tuneable lipidomes can be used as experimental chassis for screening membrane active antimicrobial peptides. Our study introduces an experimental resource and foundation for deciphering the role of lipidome complexity in membrane function and cellular fitness.

synthetic biology↗

bioRxiv2018

Body size-dependent energy storage causes Kleibers law scaling of the metabolic rate in planarians

Kleibers law, or the [3/4]-power law scaling of the metabolic rate with body mass, is considered one of the few quantitative laws in biology, yet its physiological basis remains unknown. Here, we report Kleibers law scaling in the planarian Schmidtea mediterranea. Its reversible and life history-independent changes in adult body size over 2 orders of magnitude reveal that Kleibers law does not emerge from the size-dependent decrease in cellular metabolic rate, but from a size-dependent increase in mass per cell. Through a combination of experiment and theoretical analysis of the organismal energy balance, we further show that the mass allometry is caused by body size dependent energy storage. Our results reveal the physiological origins of Kleibers law in planarians and thus have general implications for understanding a fundamental scaling law in biology.

biophysics↗

bioRxiv2018

An insulin, AMPK, and steroid hormone-mediated metabolic switch regulates the transition between growth and diapause in C. elegans

The balance between growth and quiescence depends on the global metabolic state. The dauer larva of C. elegans, a developmentally arrested stage for survival under adverse environment, undergoes a major metabolic transition. Here, we show that this switch involves the concerted activity of several regulatory pathways. Whereas the steroid hormone receptor DAF-12 controls dauer morphogenesis, the insulin pathway maintains low energy expenditure through DAF-16/FoxO, which also requires AAK-2/AMPK. DAF-12 and AAK-2 separately promote a shift in the molar ratios between competing enzymes at two key branch points within the central carbon metabolic pathway. This way, carbon atoms are diverted from the TCA cycle and directed to gluconeogenesis. When both AAK-2 and DAF-12 are suppressed, the TCA cycle is active and the developmental arrest is bypassed. Hence, the metabolic status of each developmental stage is defined by stoichiometric ratios within the constellation of metabolic enzymes and controls the transition between growth and quiescence.

biochemistry↗

bioRxiv2018

DNA-Encircled Lipid Bilayers

Lipid bilayers and lipid-associated proteins play a crucial role in biology. As in vivo studies and manipulation are inherently difficult, several membrane-mimetic systems have been developed to enable investigation of lipidic phases, lipid-protein interactions, membrane protein function and membrane structure in vitro. Controlling the size and shape, or site-specific functionalization is, however, difficult to achieve with established membrane mimetics based on membrane scaffolding proteins, polymers or peptides. In this work, we describe a route to leverage the unique programmability of DNA nanotechnology and create DNA-encircled bilayers (DEBs), which are made of multiple copies of an alkylated oligonucleotide hybridized to a single-stranded minicircle. To stabilize the hydrophobic rim of the lipid bilayer, and to prevent formation of lipid vesicles, we introduced up to 2 alkyl chains per helical that point to the inside of the toroidal DNA ring and interact with the hydrophobic side chains of the encapsulated lipid bilayer. The DEB approach described herein provides unprecedented control of size, and allows the orthogonal functionalizations and arrangement of engineered membrane nanoparticles and will become a valuable tool for biophysical investigation of lipid phases and lipid-associated proteins and complexes including structure determination of membrane proteins and pharmacological screenings of membrane proteins.

biophysics↗