Biology subjects

Modla, S.

Publications and source records attributed to Modla, S..

3 recordsLinked to original sources

bioRxiv2025

Structure of the Disulfide-rich Modules of a Striking Tandem Repeat Protein, Avian Cysteine-Rich Eggshell Membrane Protein

Avian eggshell membrane (ESM) is fabricated within the isthmus region of the oviduct and is comprised of three juxtaposed, predominantly proteinaceous layers lying between egg white and the calcified shell. The limiting membrane is less than 0.5 {micro}m in thickness and forms the osmotic barrier for the egg. This first layer provides the foundation for the successive deposition of two mats of protein fibers. Fibers from both inner and outer layers appear to have similar amino acid compositions and are notably disulfide-rich (comprising about 10% Cys). ESM has been utilized in a wide variety of applications, including nutraceutical supplements, tissue engineering, and nanofabrication, and yet fundamental questions concerning protein composition, fiber structure, and membrane assembly remain to be resolved. We previously identified an abundant disulfide-rich structural protein in chicken ESM fibers (Cysteine-rich eggshell membrane protein; CREMP) that contains multiple tandemly repeated modules. In this work, we determine a structural model for four consecutive 2-disulfide containing CREMP modules using a variety of two- and three-dimensional solution NMR experiments. CREMP modules feature an N-terminal loop region positioned above a small beta hairpin that is stabilized by a conserved pattern of disulfide bridges between Cys1-3 and Cys2-4. While the individual CREMP modules are highly ordered, the lack of long-range inter-module restraints suggests an extended structure connected by flexible linkers. Finally, the structural information obtained in this work is considered in the context of full-length CREMP proteins and compared to two other structural proteins that contain multiple tandem repeats of 2-disulfide modules. Statement of Importance and ImpactUnder the shell of an avian egg is a tough, paper-thin, three-layered protein-based structure called the eggshell membrane. This membrane has a range of medical, biomaterial and analytical applications, and yet fundamental questions concerning protein composition and assembly remain unresolved. Here, we address the 3-dimensional structure of part of a major membrane component, cysteine-rich eggshell membrane protein. The work provides new insight into the eggshell membrane and suggests an additional avenue for de-novo protein design.

biochemistry↗

bioRxiv2025

Structural Insights into Allosteric Regulation of GdpP: A Conformationally Dynamic Phosphodiesterase

The phosphodiesterase GdpP is a central regulator of the bacterial second messenger c-di-AMP and a key driver of antibiotic resistance in pathogenic Firmicutes. GdpP integrates environmental signals through its sensory PAS domain to control its C-terminal catalytic domain activity, but the molecular basis for this allosteric communication has remained unknown due to the lack of structural data for the complete cytosolic region. Here, we present the first Cryo-EM structures of the cytosolic region of Streptococcus mutans GdpP (SmGdpP74) in multiple conformational states, revealing a sophisticated tetrameric architecture that enables asymmetric catalytic regulation. Our structural and functional analyses demonstrate that SmGdpP74 operates through substrate-induced conformational transition, with the DHHA1 domain interface serving as the primary determinant of asymmetric catalysis. The non-canonical GGDEF domain functions as a tetrameric scaffolding hub that positions the DHH-DHHA1 catalytic domains, representing the first detailed example of a GGDEF domain repurposed to stabilize heterologous catalytic domains. We identify a flexible GGDEF-DHH linker as a critical coupling element that transmits conformational signals between domains, with the conserved KRSR motif acting as a molecular switch for heme-mediated inhibition. Additionally, the flexibility of this linker is essential for the enzymes catalytic activity. Based on these findings, we propose a comprehensive mechanistic model where SmGdpP74 integrates substrate availability with conformational changes for efficient hydrolysis and allosteric control of its enzymatic activity through the PAS domain. These structural insights provide a foundation for rational drug design targeting allosteric regulatory mechanisms in GdpP, potentially offering new approaches to combat antibiotic resistance in pathogenic Firmicutes.

molecular biology↗

bioRxiv2024

Chlorobaculum tepidum Outer Membrane Vesicles Are Likely a Significant Route for Biogenic Sulfur Transport.

Outer membrane-derived vesicles (OMVs) have been studied in different phyla of Gramnegative bacteria, most extensively in the Pseudomonadota, where they have been shown to participate in diverse biological and environmental processes. To date, the production of OMVs has not been reported in the Chlorobiaceae within the phylum Chlorobiota. Chlorobaculum. tepidum is the model organism for the Chlorobiaceae that synthesizes and consumes insoluble extracellular sulfur (S(0)) globules by an unknown mechanism. Here, we report evidence implicating outer membrane vesicles in biogenic S(0) globule synthesis. We demonstrate that Cba. tepidum secretes OMVs in the extracellular milieu, and that OMV concentration and size vary with growth conditions, particularly sulfide concentration. A core of 31 proteins involved in diverse biological processes such as cell wall biogenesis, inorganic ion transport and metabolism were found to be shared between OMVs, extracellular S(0) globules and Cba. tepidum intact cells. Multiple analytical methods indicated that OMVs contain S(0) and that OMVs and biogenic S(0) globules share protein and polysaccharide signatures, including lipooligosaccharides. Together these lines of evidence indicate that Cba. tepidums OMVs are one component of sulfur transport between cells and extracellular sulfur globules. IMPORTANCEAll living cells must exchange material with their environment while maintaining cellular integrity. This is a particular challenge for materials that are not water soluble, yet many bacteria utilize insoluble materials for energy conservation and as nutrients for growth. Here we show that Cba. tepidum makes outer membrane vesicles and that these vesicles are likely involved in the exchange of material with extracellular elemental sulfur globules formed and consumed by Cba. tepidum as part of its energy metabolism based on oxidizing reduced sulfur compounds like hydrogen sulfide. These data expand our basic understanding of Cba. tepidums metabolism. As elemental sulfur is an industrial by-product with a limited number of uses, the information here may help enable the use of additional sulfur compounds by Cba. tepidum to drive the synthesis of biomass and/or specialty biochemicals from waste elemental sulfur by this autotrophic bacterium.

microbiology↗