The present study identified genes and characterize hemicellulose degradation strategies of bacterial strains with high xylanolytic activity from the earthworm gut and found that each strain possesses a unique functional repertoire of genes, suggesting a variety of hemicellulolytic strategies that can be attributed to the various isoforms or different carbohydrate-binding modules of these enzymes.
ABSTRACT Polyethylene terephthalate (PET) waste represents a major environmental challenge due to limited recycling solutions. Thermophilic bacteria from geothermal environments harbor diverse enzymatic machinery adapted to extreme conditions, offering promising biocatalysts for plastic degradation; however, biological resources from Peru and other South American countries remain scarce. We characterized four bacterial strains isolated from two geothermal sites in Cajamarca, Peru, screened for PET hydrolysis at 50°C. Whole‐genome sequencing using hybrid assembly achieved near‐complete circular genomes. GTDB‐Tk classification identified three species: Neobacillus thermocopriae (strain 19A), Bacillus licheniformis (strains 16P and BI2), and Brevibacillus agri (strain BI8). Quantitative assays revealed that strain 16P achieved the highest mass loss (0.598%), followed by strain BI8 (0.449%). ATR‐FTIR analysis of the incubated sheets showed a significant reduction of the ester carbonyl index in strains 16P, 19A, and BI8 relative to both non‐incubated PET and an abiotic control, whereas strain BI2 did not differ from the controls, indicating preferential modification of ester bonds at the sheet surface. Genome mining and structure‐based homology searches identified multiple candidate enzymes similar to validated PETases and carboxylesterases, including PETase46‐like homologs in strains BI8 and 16P and a terephthalate‐active carboxylesterase homolog in strain 16P. Molecular docking supported the conservation of catalytic geometry and substrate‐binding sites in these candidates. This work represents one of the first systematic genomic and structural characterizations of putative PET‐hydrolases in Peruvian geothermal bacteria, expanding knowledge of extremophile diversity and advancing thermostable enzymes for sustainable plastic waste management.
Marco A Rivera-Jacinto, Claudia Rodríguez-Ulloa, Sara R Briones-Ramírez et al.· MicrobiologyOpen· 0 citations
A novel type III pullulan hydrolase (PulTS) was identified from the thermophilic bacterium Thermus scotoductus, representing the first bacterial member of this enzyme class. The recombinant PulTS exhibited optimal activity at pH 6.5 and 80 °C, with a specific activity of 46.3 ± 2.4 U/mg toward pullulan. The enzyme hydrolyzed pullulan into maltotriose, panose, maltose, and glucose, and also degraded soluble starch, amylose, amylopectin, dextran, glycogen, and γ-cyclodextrin. PulTS contains sequentially arranged carbohydrate-binding module family 48 (CBM48), CBM34, and glycoside hydrolase family 13 subfamily 20 (GH13_20) catalytic domains, an arrangement previously observed only in archaeal species. Deletion of CBM48 enhanced the thermostability of PulTS by approximately 3-fold, yet the enzyme still retained approximately 69% of its catalytic efficiency toward pullulan. Deletion of CBM34 resulted in no further changes. Affinity gel electrophoresis revealed that CBM48 bound pullulan and, more weakly, soluble starch, while CBM34 exhibited only weak binding ability toward pullulan. The results indicated that CBM48 was the major contributor to substrate binding and enzymatic activity, whereas CBM34 played a minor role in PulTS. This work expands our knowledge of pullulan-hydrolyzing enzymes.
Yi Gao, Zi-Jian Fu, Lan Xu et al.· International Journal of Bio...· 0 citations
This study presents the first characterization of a GH3 β-glucosidase from L. buchneri and reveals a non-classical stabilizing effect of EDTA, offering valuable insights for enzyme engineering and biocatalytic applications.
Hui Tang, Jinjian He, Can Li et al.· Metabolites· 0 citations
Penitrem A is a toxic secondary metabolite (SM) produced by
Penicillium crustosum
(
P. crustosum
) on various foods such as nuts, dairy products, and fruits. However, the biosynthetic gene clusters (BGCs) responsible for SMs including penitrem A in
P. crustosum
isolated from pears is largely unexplored.
In the current study, we performed whole-genome sequencing of
P. crustosum
KACC 411287, which can produce penitrem A and roquefortine C, and identified its SM BGCs including BGCs of the toxins. Furthermore, we conducted a comparative analysis of the penitrem A and roquefortine C BGCs against those in other fungal strains. We also analyzed the carbohydrate-active enzyme-(CAZyme-) encoding genes in
P. crustosum
KACC 411287, and compared with those of other closely related fungal strains.
The
P. crustosum
KACC 411287 genome is composed of five chromosomes, totaling approximately 32.37 Mb in size. Gene Ontology analysis using 8,520 functionally annotated proteins exhibited that the genome of
P. crustosum
KACC 411287 contains a significant abundance of genes involved in degradation of organonitrogen compounds including amino acids or carbohydrates and fungal self-protective mechanisms including SM biosynthesis. Of the 8,520 functionally annotated proteins, 546 predicted CAZymes were identified in
P. crustosum
KACC 411287. We also detected 68 SM BGCs including penitrem A and roquefortine C BGCs in
P. crustosum
KACC 411287. Furthermore, the conserved functionality analyses exhibited that each gene within the penitrem A BGC in
P. crustosum
KACC 411287 is highly conserved with the corresponding gene in four other penitrem A-producing
Penicillium
strains (above 77% amino acid sequence identity) except for
ptmH
in
P. flavigenum
IBT 14082 (20%). In contrast, the sequence identity decreased significantly (0–69% identity) in two penitrem A non-producing
Penicillium
strains.
Our data strongly indicate that the penitrem A BGCs were highly conserved among
P. crustosum
KACC 411287 and three other penitrem A-producing
Pencillium
strains. Our findings expand our knowledge about the biosynthesis of SMs including penitrem A and roquefortine C in
P. crustosum
KACC 411287 that causes blue mold rot on pears. These results could provide new insight into the biosynthesis of penitrem A and roquefortine C in
P. crustosum
KACC 411287 to find potential approaches for alleviating penitrem A or roquefortine C contamination on pears.
The study re-analyzed the draft genome of Bacillus licheniformis SMIA-2 and generated a reference-guided pseudo-scaffold. Cross-validated genome annotation identified five candidate loci associated with pectin degradation, including putative pectate lyases, polygalacturonase, and downstream uronate-catabolic genes. Submerged fermentation with passion fruit peel flour and corn steep liquor yielded crude enzymatic extracts, which were spray-dried at 110 °C using maltodextrin and microcrystalline cellulose as stabilizers. The dried formulation retained pectinase activity for 180 days at 5 °C and showed additional cellulase, amylase, xylanase, and protease activities. Pectinase displayed optimal activity at pH 8.5 and 70 °C, with stability between pH 8.0–8.5 and 65–70 °C. Despite not using a reference strain and the absence of some omics analyses, with genomic and industrial claims presented as evidence of biotechnological potential rather than definitive functional validation of individual genes, these results support a sustainable, scalable, and alkaline-tolerant enzyme platform based on agro-industrial residues.
E. Cruz, Larissa Pacheco Ferreira, A. Rosana et al.· Food Science and Biotechnolo...· 0 citations
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