Characterization and application of keratinolytic peptidases from Geobacillus sp. and Bacillus spp.

Solid keratin-rich waste management is one of essential research area in nowadays. Conventional chemical and high thermal keratin waste decomposition methods are fully explored and not enough effective for future biotechnolgy perspectives. However, traditional keratin-rich waste decomposition methods could be replaced by environmentally-friendly and economical microbial keratin waste biodegradation methods without energy wastage and essential amino acids and nutrition elements loss. Therefore, microbial bio-decomposition are attractive approach to keratin or keratin-like waste manage without any nutrition loss in eco-friendly process environment. Increased attention has been diverted to these keratinolytic peptidases because of their important potential uses in biomedicine, pharmaceutics, cosmetics and keratin waste bioconversion industries associated to the hydrolysis of keratin. Keratinolytic proteinases are next generation molecular tools for keratin hydrolysis and production of small value-added bio-active peptides. For the first time, we identified, purified and characterized native keratinolytic peptidase (BtKER) from Bacillus thuringiensis AD-12. For the first time, we created chimeric (SynKer-TT and SynKer-TM) keratinolytic peptidases with improved physicochemical properties. Eventually, we demonstrated that metabolically active secretomes from keratinolytic microorganisms can be used for eco-friendly enzymatic keratin waste management in lab-scale system

Geobacillus sp. ir Bacillus spp. sekretuojamų keratinoliziniu aktyvumu pasižyminčių peptidazių charakterizavimas ir pritaikymas

Solid keratin-rich waste management is one of essential research area in nowadays. Conventional chemical and high thermal keratin waste decomposition methods are fully explored and not enough effective for future biotechnolgy perspectives. However, traditional keratin-rich waste decomposition methods could be replaced by environmentally-friendly and economical microbial keratin waste biodegradation methods without energy wastage and essential amino acids and nutrition elements loss. Therefore, microbial bio-decomposition are attractive approach to keratin or keratin-like waste manage without any nutrition loss in eco-friendly process environment. Increased attention has been diverted to these keratinolytic peptidases because of their important potential uses in biomedicine, pharmaceutics, cosmetics and keratin waste bioconversion industries associated to the hydrolysis of keratin. Keratinolytic proteinases are next generation molecular tools for keratin hydrolysis and production of small value-added bio-active peptides. For the first time, we identified, purified and characterized native keratinolytic peptidase (BtKER) from Bacillus thuringiensis AD-12. For the first time, we created chimeric (SynKer-TT and SynKer-TM) keratinolytic peptidases with improved physicochemical properties. Eventually, we demonstrated that metabolically active secretomes from keratinolytic microorganisms can be used for eco-friendly enzymatic keratin waste management in lab-scale system

Keratinous waste decomposition and peptide production by keratinase from Geobacillus stearothermophilus AD-11.

A keratinolytic proteinase was cloned from thermophilic bacterium Geobacillus stearothermophilus AD-11 and was expressed in Escherichia coli BL21(DE3). Recombinant keratinolytic proteinase (RecGEOker) with an estimated molecular weight of 57 kDa was purified and keratinase activity was measured. RecGEOker showed optimal activity at pH 9 and 60 °C. Recombinant keratinolytic proteinase showed the highest substrate specificity toward keratin from wool > collagen > sodium caseinate > gelatin > and BSA in descending order. RecGEOker is applicable for efficient keratin waste biodegradation and can replace conventional non-biological hydrolysis processes. High-value small peptides obtained from enzymatic biodegradation by RecGEOker are suitable for industrial application in white and/or green biotechnology for use as major additives in various products

Optimizing the growth conditions of the selected plant-growth-promoting rhizobacteria Paenibacillus sp. MVY-024 for industrial scale production

In this study, thirteen isolates, which were possibly expected to fix nitrogen, were isolated from soil and pea root nodules and identified by the gene analysis of 16S rDNA sequences. Two of these isolates that were able to form endospores and grow on nitrogen-free media were selected for spring wheat development research. The isolate Paenibacillus sp. S7 identified as Paenibacillus polymyxa was found to significantly increase the amount of ammonium and mineral N amounts in the soil. Furthermore, increased nitrogen accumulation in grains and a chlorophyll index were obtained after wheat treatment. Paenibacillus sp. S7 isolate was selected for further studies and the accession number MT900581 and strain name MVY-024 in NCBI nucleotide bank for this isolate were assigned. During the cultivation of Paenibacillus sp. MVY-024, sugarcane molasses and a yeast extract were determined as the most suitable carbon and nitrogen sources, whose optimal concentrations were 100 g L−1 and 10 g L−1, respectively. The optimal pH range for the cell culture was between 6.5 and 7.0, and the optimal air flow rate was 0.4 vvm. It was found that the air flow has an effect on biomass production and endospore formation. After Paenibacillus sp. MVY-024 biomass cultivation optimization, the cultured cell number was, on average, 2.2 × 109 cfu m L−1

Phosphate Solubilizing Microorganism Bacillus sp. MVY-004 and Its Significance for Biomineral Fertilizers’ Development in Agrobiotechnology

In this study, a phosphate solubilizing microorganism was isolated from the soil of an agricultural field in Lithuania. Based on 16S rRNA gene sequence analysis, the strain was identified as Bacillus sp. and submitted to the NCBI database, Sector of Applied Bio-catalysis, University Institute of Biotechnology, Vilnius, Lithuania and allocated the accession number KY882273. The Bacillus sp. was assigned with the number MVY-004. The culture nutrient medium and growth conditions were optimized: molasses was used as a carbon source; yeast extract powder was used as an organic source; NH4H2PO4 was used as a nitrogen source; the culture growth temperature was 30 ± 0.5 °C; the initial value of pH was 7.0 ± 0.5; the partial pressure of oxygen (pO2) was 60 ± 2.0; the mixer revolutions per minute (RPM) were 25–850, and the incubation and the fermentation time was 48–50 h. Analysis using Liquid Chromatography Time-of-Flight Mass Spectrometry (LC-TOF/MS) results showed that Bacillus sp. MVY-004 produced organic acids such as citric, succinic, 2-ketogluconic, gluconic, malic, lactic, and oxalic acids. Furthermore, the experiment showed that Bacillus sp. MVY-004 can also produce the following phytohormones: indole-3-acetic (IAA), jasmonic (JA), and gibberellic (GA3) acids. In the climate chamber, the experiment was performed using mineral fertilizer (NPS-12:40:10 80 Kg ha−1) and mineral fertilizers in combination with Bacillus sp. MVY-004 cells (NPS-12:40:10 80 Kg ha−1 + Bacillus sp. MVY-004) in loamy soil. Analysis was performed in three climate conditions: normal (T = 20 °C; relative humidity 60%); hot and dry (T = 30 °C; relative humidity 30%); hot and humid (T = 30 °C; relative humidity 80%)

Novel leaderless bacteriocin geobacillin 6 from thermophilic bacterium Parageobacillus thermoglucosidasius

Bacterial resistance to conventional antibiotics has urged us to develop alternative strategies against bacterial pathogens. Moreover, a demand for food products containing no chemical preservatives has led us to search for new alternative technologies for food preservation. Bacteriocins – ribosomally synthesized antimicrobial peptides – have been proposed as a new alternative to conventional antibiotics or chemicals for food preservation. This study describes biosynthesis and characterization of a novel leaderless bacteriocin, geobacillin 6, which was identified in the thermophilic bacterium Parageobacillus thermoglucosidasius. Its amino acid sequence shows low similarity to other bacteriocins and it is the first leaderless-type bacteriocin identified in thermophilic bacteria. Based on structure assessment, the bacteriocin forms a multi-helix bundle. Geobacillin 6 exhibits a relatively narrow antimicrobial spectrum, it is active in the μM range and against Gram-positive bacteria, mostly thermophilic species closely related to the producer strain. Bacteriocin demonstrates stability over pH 3–11 and is highly thermostable, retaining 100% of its activity after incubation at 95°C for 6 h. Geobacillin 6 has potential in the food industry and biotechnological processes where contamination with thermophilic bacteria is undesirable

The Co-Inoculation Effect on <i>Triticum aestivum</i> Growth with Synthetic Microbial Communities (SynComs) and Their Potential in Agrobiotechnology

The use of rhizospheric SynComs can be a new and sustainable strategy in the agrobiotechnology sector. The objective of this study was to create the most appropriate SynCom composition; examine the ability to dissolve natural rock phosphate (RP) from Morocco in liquid-modified NBRIP medium; determine organic acids, and phytohormones; and verify plant growth promoting and nutrition uptake effect in the pot experiments of winter wheat (Triticum aestivum). A total of nine different microorganisms were isolated, which belonged to three different genera: Bacillus, Pseudomonas, and Streptomyces. Out of the 21 treatments tested, four SynComs had the best phosphate-dissolving properties: IJAK-27+44+91 (129.17 mg L−1), IIBEI-32+40 (90.95 µg mL−1), IIIDEG-45+41 (122.78 mg L−1), and IIIDEG-45+41+72 (120.78 mg L−1). We demonstrate that these SynComs are capable of producing lactic, acetic, gluconic, malic, oxalic, citric acids, and phytohormones such as indole-3-acetic acid, zeatin, gibberellic acid, and abscisic acid. In pot experiments with winter wheat, we also demonstrated that the designed SynComs were able to effectively colonize the plant root rhizosphere and contributed to more abundant plant growth characteristics and nutrient uptake as uninoculated treatment or uninoculated treatment with superphosphate (NPK 0-19-0). The obtained results show that the SynCom compositions of IJAK-27+44+91, IIBEI-32+40, IIIDEG-45+41, and IIIDEG-45+41+72 can be considered as promising candidates for developing biofertilizers to facilitate P absorption and increase plant nutrition