Biology subjects

Bleicher, L.

Publications and source records attributed to Bleicher, L..

3 recordsLinked to original sources

bioRxiv2026

Carbohydrate-active enzymes of giant viruses: Molecular and biochemical characterization of glycosyl hydrolases from algae-infecting chloroviruses

Microbial hydrolases are considered to be promising enzymes for pathogen control. Bacterial and viral chitinases of the glycosyl hydrolase (GH) 18 family are important biological macromolecules with antifungal and anti-insect activity. Chloroviruses, nucleocytoplasmic large DNA viruses (NCLDVs) that infect unicellular green algae have a considerable number of genes involved in carbohydrate metabolism, including the chitinase GH18 family. In this study, we investigated the abundance and diversity of chitinases in chlorovirus genomes using a combination of silico and in vitro strategies, and characterized these enzymes at a molecular and biochemical level. Different enzymatic profiles were observed in Chlorovirus subgenera revealing the different viral machinery related to host species. We performed a comprehensive biochemical characterization of three heterologous expressed GH18 domains, which revealed their endo and exochitinase activity and thermostability. Crystallographic analysis of the GH18 domain by X-ray diffraction yielded a structure at 1.0 [A] resolution, representing the highest-resolution structure reported to date for a giant viral protein and showing lower predominancy of residue coevolution compared GH18 chitinases from other organisms. Additionally, our binding site characterization predicted high conservation in betachloroviruses and gammachloroviruses, and less so in alphachloroviruses. Lastly, these enzymes did not inhibit fungal growth of medical and agricultural importance species in vitro but exhibited high inhibitory activity against different algae at nanogram/mL range. Together, our experimental and computational data show that evolutionary events may contribute to maintaining viral chitinases enzymatic activity and specificity. These findings highlight the potential of virus-derived enzymes as promising new biotechnological tools for microbial control against different algal strains.

microbiology↗

bioRxiv2026

Structural and functional insights into yeast Rqc1p, a protein required for thermotolerance with potential nuclear localization

Protein homeostasis - i.e., proteostasis - is the biological process by which the qualitative and quantitative balance of the proteome is conducted, either by preserving functionally relevant proteins or by degrading unnecessary ones. Stress conditions can modulate cellular proteostasis in order to promote cytoprotection and preserve the viability of living organisms. Among the cellular pathways already described that can play an important role in preserving biological functions by modulating proteostasis are the heat shock response and the ribosome quality control pathways. In this work, we show that the Rqc1p protein is necessary for the thermoadaptation of S. cerevisiae to heat shock, as RQC1-deficient yeast is sensitive to elevated temperatures. In silico approaches - such as multiple sequence alignment, structural analysis, and molecular dynamics simulations - confirmed earlier predictions that Rqc1p shares characteristics with the bHLH family of proteins. We also verified, through computational prediction of sub-cellular localization, that S. cerevisiae Rqc1p contains nuclear localization signals, suggesting that this protein can potentially be translocated toward the nucleus, thereby broadening its current range of recognized biological functions in this organism. Also, analysis of yeast transcriptomes subjected to heat shock showed that Rqc1p mRNA levels do not fluctuate in response to heat shock, suggesting that cellular concentrations of Rqc1p are already at optimal levels to elicit a rapid and effective response during thermal stress in S. cerevisiae.

biochemistry↗

bioRxiv2021

Resurrecting Golgi proteins to grasp Golgi ribbon formation and self-association under stress

The Golgi complex is a membranous organelle located in the heart of the eukaryotic secretory pathway. A subfamily of the Golgi matrix proteins, called GRASPs, are key players in the stress-induced unconventional secretion, the Golgi dynamics during mitosis/apoptosis, and Golgi ribbon formation. The Golgi ribbon is vertebrate-specific and correlates with the appearance of two GRASP paralogs (GRASP55/GRASP65) and two coiled-coil Golgins (GM130/Golgin45), which interact with each other in vivo. Although essential for the Golgi ribbon formation and the increase in Golgi structural complexity, the molecular details leading to their appearance only in this subphylum are still unknown. Moreover, despite the new functionalities supported by the GRASP paralogy, little is known about the structural and evolutionary differences between these paralogues. In this context, we used ancestor sequence reconstruction and several biophysical/biochemical approaches to assess the evolution of the GRASP structure, flexibility, and how they started anchoring their Golgin partners. Our data showed that the Golgins appeared in evolution and were anchored by the single GRASP ancestor before gorasp gene duplication and divergence in Metazoans. After the gorasp divergence, variations inside the GRASP binding pocket determined which paralogue would recruit each Golgin partner (GRASP55 with Golgin45 and GRASP65 with GM130). These interactions are responsible for the proteins specific Golgi locations and the appearance of the Golgi ribbon. We also suggest that the capacity of GRASPs to form supramolecular structures is a long-standing feature, which likely affects GRASPs participation as a trigger of the stress-induced secretory pathway.

biophysics↗