Non-edible cellulose-containing biomass is a promising and abundant feedstock for simple sugar production. This study presents the results of different cellulose-containing materials (CCM) hydrolysis experiments with P. verruculosum enzyme complexes in laboratory conditions. Among the non-pretreated substrates, only a few had a relatively high convertibility—soy bean husks (31%) and sugar beat pulp (20%)—while wheat straw, oat husks, sunflower peals, and corn stalks had a low convertibility of 3% to 12%. This indicates that a major part of CCM needs pretreatment. Steam-exploded (with Ca(OH)₂) soy bean and oat husks (76% and 58%), fine ball-milled aspen wood and nitric acid-pretreated aspen wood (62% and 78%), and steam-exploded (with sulfuric acid) corn stalks (55%) had a high convertibility. Woody biomass pretreated with pulp and paper mills also had a high convertibility (56–78%)—e.g., never dried kraft hardwood and softwood pulp (both bleached and unbleached). These results demonstrate that effective cellulose-containing material processing into simple sugars is possible. Simple sugars derived from CCM using P. verruculosum preparation are a promising feedstock for the microbiological production of biofuels (bioethanol and biobutanol), aminoacids, and organic acids (e.g., lactic acid for polylactic acid production).

Pancreatic ductal adenocarcinoma (PDAC) is a highly fatal malignancy that has the worst 5-year survival rate of all of the common malignant tumors. Surgery, chemotherapy, and/or chemoradiation remain the main tactics for PDAC treatment. The efficacy of chemotherapy is often compromised because of the substantial risk of severe toxicities. In our study, we focused on identification of polymorphisms in the genes involved in drug metabolism, DNA repair and replication that are associated with inter-individual differences in drug-induced toxicities. Using the microarray, we genotyped 12 polymorphisms in the DPYD, XPC, GSTP1, MTHFR, ERCC1, UGT1A1, and TYMS genes in 78 PDAC patients treated with FOLFIRINOX. It was found that the TYMS rs11280056 polymorphism (6 bp-deletion in TYMS 3′-UTR) predicted grade 1–2 neurotoxicity (p = 0.0072 and p = 0.0019, according to co-dominant (CDM) and recessive model (RM), respectively). It is the first report on the association between TYMS rs11280056 and peripheral neuropathy. We also found that PDAC patients carrying the GSTP1 rs1695 GG genotype had a decreased risk for grade 3–4 hematological toxicity as compared to those with the AA or AG genotypes (p = 0.032 and p = 0.014, CDM and RM, respectively). Due to relatively high p-values, we consider that the impact of GSTP1 rs1695 requires further investigation in a larger sample size.

Sugar beet pulp is a byproduct of white sugar production, and it is quite significant in terms of volume. Every year, tens of millions of tons of beet pulp are produced around the world. However, only a fraction of it is currently used, mainly as animal feed. The composition of beet pulp includes plant polysaccharides, such as cellulose, arabinan, and pectin. Through the process of enzymatic hydrolysis, these polysaccharides are converted into technical C6/C5 sugars, which can be further used as a substrate for the microbial synthesis of various substances, including biofuels, organic acids, and other green chemistry molecules. The current study was designed with a primary objective that focused on the development of a strain that had the potential for enhanced productivity and the capacity to produce enzymes suitable for beet pulp hydrolysis. The pelA and abfA genes, which encode pectin lyase and arabinofuranosidase, respectively, in the fungus Penicillium canescens (VKPM F-178), were cloned and successfully expressed in the recipient strain Penicillium verruculosum B1-537 (VKPM F-3972D). New recombinant strains were created using the expression system of the mycelial fungus P. verruculosum B1-537, which is capable of simultaneously producing pectin lyase and arabinofuranosidase, as well as homologous cellulases. The screening of strains for increased enzymatic activity towards citrus pectin, sugar beet branched arabinan, and microcrystalline cellulose revealed that a B4 clone of P. verruculosum exhibited the greatest potential in sugar beet pulp cake hydrolysis. This clone was selected as the basis for the creation of a new enzyme preparation with enhanced pectin lyase, arabinase, and cellulase activities. The component composition of the enzyme preparation was determined, and the results indicated that the enzyme content comprised approximately 11% pectin lyase, 40% arabinofuranosidase, and 40% cellulases. The primary products of the enzymatic hydrolysis of the unpretreated beet pulp cake were arabinose and glucose. The degree of arabinan and cellulose conversion was observed to be up to 50% and 80%, respectively, after a period of 48 to 72 h of hydrolysis. The new B4 preparation was observed to be highly efficacious in the hydrolysis of beet cake at elevated concentrations of solids (up to 300 g/L) within the reaction mixture. The newly developed strain, as a producer of pectin lyase, arabinofuranosidase, and cellulase complexes, has the potential to be utilized for the bioconversion of sugar beet processing wastes and for the efficient generation of highly concentrated solutions of technical sugars for further implementation in processes of microbial synthesis.

Filamentous fungi are widely used in biotechnological processes because they secrete significant amounts of protein, use inexpensive nutrient media, and are predictably scalable in technological processes. Penicillium verruculosum B1-537 (now renamed Talaromyces verruculosus) produces large amounts of secreted protein (up to 70 g/L) and is used for large-scale enzyme production. Although P. verruculosum has an excellent protein expression system under the control of a strong cbh1 promoter, some heterologous enzymes such as Aspergillus awamori glucoamylase (aaGlaA) are still produced in insufficient quantities (15–20% of the total secreted protein), and this limits the application of enzyme preparations derived from P. verruculosum strains in the alcohol industry for the enzymatic treatment of grain starch together with α-amylase. One of the well-known approaches to addressing this is signal peptide replacement to increase protein expression. Therefore, the aim of this study was to investigate the effectiveness of signal peptide replacement. Various signal peptides (SPs), which were previously used for other well-expressed heterologous proteins, such as xylanases, β-glucosidases, and others, were analyzed to determine their effect on aaGlaA secretion. Five plasmids containing signal peptide sequences fused to the glaA gene were constructed and used to transform P. verruculosum. The resulting strains were cultured and screened for protein content and glucoamylase activity. Copy number analysis was performed on the most productive strains. The best one was an SP of homologous glucoamylase in P. verruculosum (pvGlaA). The use of this particular SP increased the secretion of heterologous aaGlaA by 2.5 times when cultivating recombinant strains on cellulose-containing fermentation media for P. verruculosum. Thus, SP replacement is a useful way to increase expression levels in the P. verruculosum expression system. Application of this method in P. verruculosum could address some productivity issues and enable the large-scale production of other industrial and food enzymes.

Aim. To assess the association between the carriage of minor allelic variants of 8 genes that encode key enzymes involved in the metabolism of anticancer drugs (DPYD, GSTP1, MTHFR, UGT1A1) and cell repair (XPC, ERCC1, TYMP, NQO1) and the severity of adverse drug events in patients with common gastrointestinal tumors. Tasks. To study the frequency of minor allelic variants of the DPYD, GSTP1, MTHFR, UGT1A1, XPC, ERCC1, TYMP, NQO1 genes; to assess the frequency and severity of adverse drug events of chemotherapy treatment in the study population. Materials and methods. For the period from October 2020 to April 2021, 56 patients (women 29, men 27) with verified malignant tumors of the gastrointestinal tract were included in a prospective clinical study as a part of the RSF grant No. 20-75-10158. The mean age was 62.311.4 years. Colon cancer was detected in 24 patients, tumors of the esophagus and stomach in 19 patients, tumors of pancreas and biliary tract in 13 patients. First-line palliative chemotherapy was given to 27 patients, adjuvant 19 patients, neoadjuvant 10 patients. All patients had not previously received cytotoxic or radiation treatment. Point nucleotide variants of genes DPYD, XPC, GSTP1, MTHFR, ERCC1, UGT1A1, TYMPS, NQO1 were determined by hybridization analysis on biological microchips. Differences in the tolerance of cytotoxic therapy (5-fluorouracil, platinum preparations, irinotecan) depending on the genotype were assessed using Fishers exact test. Results. The average number of chemotherapy courses received was 4.22.6 (112). There was a statistically significant difference in the tolerability of chemotherapy in patients with minor allelic variants of the GSTP1 rs1695 (p=0.03), ERCC1 rs11615 (p=0.01), and UGT1A1 rs8175347 (p=0.003) genes. Conclusion. The use of hybridization analysis on biological microchips to assess allelic variants responsible for the tolerability of cytotoxic therapy is reasonable and requires further prospective assessment.