bioRxiv Science⌕ Search
Home/Archive/bioRxiv

bioRxiv · 10.1101/2022.10.18.512808

Profiling of the β-glucosidases identified in the genome of Penicillium funiculosum: Insights from genomics, transcriptomics, proteomics and homology modelling studies

Okereke, O. E.·Gupta, M.·Ogunyewo, O. A.·Sharma, K.·Yazdani, S. S.
View original source ↗Export metadata ↓

Abstract

Enzymatic lignocellulosic biomass conversion to bioethanol is dependent on efficient enzyme systems with {beta}-glucosidase as a key component. In this study, we performed in-depth profiling of the various {beta}-glucosidases present in the genome of the hypercellulolytic fungus; Penicillium funiculosum using genomics, transcriptomics, proteomics and molecular dynamics simulation approaches. Of the eight {beta}-glucosidase genes identified in the P. funiculosum genome, three were found to be extracellular, as evidenced by presence of signal peptides and mass spectrometry. Among the three secreted {beta}-glucosidase, two belonged to the GH3 and one belonged to GH1 families. Modelled structures of these proteins predicted a deep and narrow active site for the GH3 {beta}-glucosidases (PfBgl3A and PfBgl3B) and a shallow open active site for the GH1 {beta}-glucosidase (PfBgl1A). The enzymatic assays indicated that P. funiculosum secretome showed high {beta}-glucosidase activities with prominent bands on 4-methylumbelliferyl {beta}-D-glucopyranoside (MUG) zymogram. To understand the contributory effect of each of the three secreted {beta}-glucosidases (PfBgls), the corresponding gene was deleted separately and the effect of the deletion on {beta}-glucosidase activity of the secretome was examined. Although not the most abundant {beta}-glucosidase, PfBgl3A was found to be the most significant one as evidenced by a 42 % reduction in {beta}-glucosidase activity in the {Delta}PfBgl3A strain. To improve the thermostability, two mutants of PfBgl3A were designed with the help of molecular dynamics (MD) simulation and were expressed in Pichia pastoris for evaluation. The PfBgl3A mutant (Mutant A) gave 1.4 fold increase in the half-life (T1/2) of the enzyme at 50{degrees}C. IMPORTANCECommercially available cellulases are majorly produced from Trichoderma reesei. However, external supplementation of the cellulase cocktail from this host with exogenous {beta}-glucosidase is often required to achieve desired optimal saccharification of cellulosic feedstocks. This challenge has led to exploration of other cellulase-producing strains because of the importance of this class of enzymes in the cellulose deconstruction machinery. The non-model hypercellulolytic fungus Penicillium funiculosum has been studied in recent times and identified as a promising source of industrial cellulases. Various genetic interventions targeted at strain improvement for cellulase production have been performed. However, the {beta}-glucosidases of this strain have remained largely understudied. This study, therefore, reports profiling of all the eight {beta}-glucosidases of P. funiculosum via molecular and computational approaches and enhancing thermostability of the most promising {beta}-glucosidase via protein engineering. The results of this study set the background for future engineering strategies to transform the fungus into an industrial workhorse.

Explore related subjects

microbiologyPreprint

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Okereke, O. E., Gupta, M., Ogunyewo, O. A., Sharma, K., Yazdani, S. S.. 2022-10-19. Profiling of the β-glucosidases identified in the genome of Penicillium funiculosum: Insights from genomics, transcriptomics, proteomics and homology modelling studies. https://doi.org/10.1101/2022.10.18.512808

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

A population-scale landscape of the subgingival microbiome reveals divergent routes to periodontal dysbiosis

Periodontitis is an archetypical mucosal inflammatory disease in which microbiome dysbiosis at the tooth-epithelial interface interacts with host genetic and behavioral risk factors to drive immune-mediated tissue destruction. Although subgingival microbiome compositional shifts are thought to parallel disease severity, microbiome variation at the population-level and its relationship to periodontal clinical phenotypes and disease-modifying factors remain poorly defined. Here, we use unsupervised manifold learning to map the compositional landscape of the subgingival microbiome in 1,355 adults spanning periodontal health to severe periodontitis. We identified eight latent microbiome states organized along a branching continuum from eubiosis to dysbiosis. An intermediate microbial configuration marked ecological destabilization and bifurcation into two distinct periodontitis-associated dysbiotic trajectories, distinguished by links to gingival inflammation and smoking. Although the microbiome trajectories broadly tracked periodontal destruction, a minority of individuals showed discordant microbiome-clinical phenotypes, with some individuals with periodontitis retaining otherwise eubiotic microbiomes enriched for low-abundance pathobionts, while some cases of health or mild disease had highly dysbiotic communities, suggesting distinct host susceptibility. Together, these findings define a population-scale ecological landscape of the subgingival microbiome, reveal divergent trajectories to periodontal dysbiosis, and highlight heterogeneity in the relationship between microbial community structure and clinical disease expression.

microbiology↗
bioRxiv2026

The iron-binding siderophore enterobactin is required for the response of multi-drug resistant Klebsiella pneumoniae to zinc limitation

To persist during infection Klebsiella pneumoniae must overcome nutrient iron and zinc limitation imposed by the host immune system through a process called nutritional immunity. Secreted small molecule siderophores are a major virulence determinant of Klebsiella pneumoniae pathogenesis and are presumed to overcome nutritional immunity by binding iron for bacterial acquisition. In this work, we set out to identify how a multi-drug resistant K. pneumoniae grows in zinc limited environments. Using unbiased transcriptomics, proteomics, and an arrayed transposon screen, we identified that synthesis and uptake of the siderophore enterobactin is required to allow for growth in low zinc conditions. Iron-specific chelators did not replicate this phenotype and addition of supplemental iron through heme in growth media could not complement severe growth defects of enterobactin mutant K. pneumoniae experiencing zinc limitation. Finally, zinc starvation induced enterobactin production independent of the canonical zinc uptake regulator (Zur) transcription factor suggesting an unidentified regulatory mechanism by which Gram-negative pathogens may respond to zinc stress. Together, these studies expand the role of enterobactin beyond iron regulation and highlight a previously unreported link between iron and zinc homeostasis in Klebsiella pneumoniae.

microbiology↗
bioRxiv2026

A microbiota-derived protease links phage susceptibility to host epithelial responses

Bacteriophages are major ecological drivers of gut microbial ecology, yet whether bacterial mechanisms that determine phage susceptibility have consequences for the mammalian host remains poorly understood. Here, we identify dipeptidyl peptidase 11 (Dpp11a), the predominant active serine protease of the prevalent gut commensal Phocaeicola vulgatus, as an unexpected bacterial defence factor. Dpp11a protects against environmental proteases and confers resistance to bacteriophage infection. Metatranscriptomic analyses further reveal increased expression of both dpp11a and P. vulgatus-associated phage transcripts in ulcerative colitis stool samples, indicating that both components of this interaction are transcriptionally active in disease-associated human microbiomes. Using the microfluidic gut-on-a-chip co-culture model HuMiX, we show that the absence of Dpp11 is accompanied by altered epithelial tight-junction remodelling during phage-bacterial infection. Together, our findings reveal that the consequences of bacterial phage defence can extend beyond phage-bacterium interactions to the mammalian epithelium.

microbiology↗