Biology subjects

Noteborn, W. E. M.

Publications and source records attributed to Noteborn, W. E. M..

3 recordsLinked to original sources

bioRxiv2025

Using cryogenic electron microscopy methods to gain insight into structure and initial host attachment of the flagellotropic bacteriophage 7-7-1

Understanding the structural and functional mechanisms of bacteriophage 7-7-1, the flagellotropic phage infecting Agrobacterium sp. H13-3, offers promising insights into phage-host interactions. Using single particle analysis (SPA) cryo-electron microscopy (cryo-EM), we determined the capsid and tail structure, and built atomic models of capsid hexamers, pentamers and tail. Combined with cryo-electron tomography (cryo-ET) and machine learning methodologies, our findings indicate that phage 7-7-1 uses capsid fibers to establish initial contact with the host flagellum, followed by subsequent attachment to cell surface receptors. Proteinase K treatment confirmed the time-dependent degradation of capsid fibers. The study also demonstrated that capsid fibers are flexible and can interact with other phages and host flagella, suggesting a cooperative infection strategy. These results provide crucial structural insights and may open avenues for developing phage-based therapeutics against resistant bacterial pathogens.

microbiology↗

bioRxiv2024

Promiscuous NAD-dependent dehydrogenases enable efficient bacterial growth on the PET monomer ethylene glycol

Ethylene glycol is widely used as antifreeze agent and monomer of the ubiquitous plastic PET (polyethylene terephthalate). Its global production amounts to more than 50 million tons per year, and it constitutes an environmental pollutant of increasing concern. Although it is generally accepted that bacteria oxidize ethylene glycol to use it as growth substrate, the enzymes involved in this process are not well understood. Here we show that the soil bacterium Paracoccus denitrificans is able to assimilate ethylene glycol efficiently via NAD-dependent alcohol and aldehyde dehydrogenases. Using comparative proteome analysis, we identify a previously unknown gene cluster that is strongly expressed in the presence of ethylene glycol. We report the kinetic parameters and cryo-EM structures of EtgB and EtgA, the key enzymes encoded by this etg gene cluster. These novel biocatalysts pave the way for more efficient biotechnological conversion of ethylene glycol. We furthermore show that the transcriptional activator EtgR controls expression of the etg gene cluster. Directed evolution of P. denitrificans on ethylene glycol results in faster growing strains, which is enabled by increased activities of EtgB and EtgA. Bioinformatic analysis reveals that the etg gene cluster and variants thereof are widely distributed among Proteobacteria, suggesting a previously underappreciated role of NAD-dependent dehydrogenases in microbial ethylene glycol assimilation.

microbiology↗

bioRxiv2023

Super-resolution fluorescence imaging of cryosamples does not limit achievable resolution in cryoEM

Correlated super-resolution cryo-fluorescence and cryo-electron microscopy (cryoEM) has been gaining popularity as a method to investigate biological samples with high resolution and specificity. A concern in this combined method (called SR-cryoCLEM), however, is whether and how fluorescence imaging prior to cryoEM acquisition is detrimental to sample integrity. In this report, we investigated the effect of high-dose laser light irradiation on apoferritin samples prepared for cryoEM with excitation wavelengths commonly used in fluorescence microscopy, and comparing these samples to controls that were kept in the dark. We found that laser illumination, of equal duration and intensity as used in super-resolution cryomicroscopy and in the presence of high concentrations of fluorescent protein, did not affect the achievable resolution in cryoEM, with final reconstructions reaching resolutions of ~1.8 [A] regardless of the illumination conditions. The finding that super-resolution fluorescence imaging of cryosamples prior to cryoEM data acquisition does not limit the achievable resolution suggests that super-resolution cryo-fluorescence microscopy and in situ structural biology using cryoEM are entirely compatible. Graphical abstract O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY

molecular biology↗