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Cervical cancer accounts for about two-thirds of all cancer cases linked etiologically to Human Papilloma Virus (HPV). 15 oncogenic HPV types can cause cervical cancer, of which HPV16 and HPV18 combinedly account for about 70% of it. So, effective epitope design for the clinically relevant HPV types 16 and 18 would be of major medical benefit. Here, a comprehensive analysis is carried out to predict the epitopes against HPV types 16 and 18 through “reverse vaccinology” approach. We attempted to identify the evolutionarily conserved regions of major capsid protein (L1) as well as minor capsid protein (L2) of HPV and designed epitopes within these regions. In this study, we analyzed about 49 and 27 sequences of HPV L2 and L1 proteins respectively. Since we found that the intertype variability of L2 is higher than for L1 proteins, our analysis was emphasized on epitopes of L1 of HPV types 16 and 18. We had selected HLA-A*0201, DRB1*1501, DQB1*0602, DRB1*0401 and DQB1*0301 alleles for the prediction of T cell epitopes of L1 of HPV 16 and 18. Finally, we reported that predicted epitope sequences EEYDLQFIFQLCKITLTA, and RHGEEYDLQFIFQLCKITLTA of L1 protein of HPV 16, and LPDPNKF, PETQRLVWAC, PVPGQYDA, YNPETQRLVWAC, DTGYGAMD, PVPGQYDATK, KQDIPKVSAYQYRVFRV, RDNVSVDYKQTQLCI and YSRHVEEYDLQFIF of L1 protein of HPV 18 could be therapeutic tools for vaccine design against HPV.

Biophysical understanding of amorphous protein aggregation can significantly impact diverse area of biotechnology. Here, we report the time dependent salt-induced formation of amorphous aggregation as monitored by fluorescence self-quenching and compare the results with conventional methods for detecting protein aggregation [static light scattering (LS) and dynamic light scattering (DLS)]. As a model protein, we used a bovine pancreatic trypsin inhibitor (BPTI) variant extended by two glycines (C2G) at its C terminus, and three variants where three types of Solubility Controlling Peptide tags (SCP tags) made of five serines (C5S), alanines (C5A) or aspartic acids (C5D) were added to the C terminus of C2G. All variants have a native-like BPTI structure and trypsin inhibitory activity, but different solubilities controlled by the SCP tags. The BPTIs were labeled using NHS-Fluorescein (FAM) conjugated to BPTI's lysines, and we measured the changes in fluorescence intensity occurring upon the addition of NaCl. The fluorescence of all FAM-BPTIs decreased almost immediately, albeit to a different extent, upon addition of salt and became constant after 10 min for 24 h or more. On the other hand, LS and DLS signal changes were dependent on the type of tags. Namely, C2G's LS and DLS signals changed immediately, the signals of C5S and C5A tagged FAM-BPTIs increased slowly from 10 min to 24 h, and those of C5D remained constant. These observations indicated the presence of at least one intermediate step, with increased protein-protein interaction yielding a 'molecular condensation' phase. According to this model, C2G would rapidly turn from 'condensates' to aggregates, whereas C5S and C5A tagged FAM-BPTIs would do so slowly, and the soluble C5D tagged variant would remain in the molecular condensation state.

A commercial plant probiotic product was developed employing Bacillus subtilis CW-S in submerged fermentation. The effects of molasses and urea on cell growth were investigated with the goal of low-cost manufacturing. Plackett–Burman and Central-Composite Design (CCD) were utilized to optimize production parameters to maximize productivity. The stability of the formulated product and its efficacy in cultivating minituber in aeroponics and industrial-grade potatoes in the field were assessed. The results showed that the medium BS10 (molasses and urea) produced satisfactory cell density (7.19 × 108 CFU/mL) as compared to the control (1.51 × 107 CFU/mL) and BS1-BS9 (expensive) media (1.84 × 107–1.37 × 109 CFU/mL). According to validated CCD results, optimized parameters fitted well in pilot (300 L; 2.05 × 109 CFU/mL) and industrial (3000 L; 2.01 × 109 CFU/mL) bioreactors, resulting in a two-fold increase in cell concentration over laboratory (9.84 × 108 CFU/mL) bioreactors. In aeroponics, CW-S produced excellent results, with a significant increase in the quantity and weight of minitubers and the survival rate of transplanted plantlets. In a field test, the yield of industrial-grade (> 55 mm) potatoes was increased with a reduction in fertilizer dose. Overall, the findings suggest that CW-S can be produced commercially utilizing the newly developed media and optimized conditions, making plant probiotics more cost-effective and accessible to farmers for crop cultivation, particularly in aeroponic minituber and industrial-grade potato production.

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