RESUMO
Chitinous fishery by-products have great application in the production of various bioactive compounds. In this study, Paenibacillus elgii TKU051, a protease-producing bacterial strain, was isolated using a medium containing 1% squid pens powder (SPP) as the sole carbon/nitrogen (C/N) source. P. elgii TKU051 was found to produce at least four proteases with molecular weights of 100 kDa, 57 kDa, 43 kDa, and 34 kDa (determined by the gelatin zymography method). A P. elgii TkU051 crude enzyme cocktail was optimally active at pH 6-7 and 60 °C. The 2,2-diphenyl-1-picrylhydrazyl radical scavenging activity and α-glucosidase inhibitory activity of the hydrolysates obtained from the hydrolysis of shrimp shell powder, shrimp head powder, shrimp meat powder, fish head powder and soya bean powder catalyzed by the P. elgii TkU051 crude enzyme cocktail were also evaluated. P. elgii TKU051 exhibited a high deproteinization capacity (over 94%) on different kinds of shrimp waste (shrimp heads and shells; fresh and cooked shrimp waste; shrimp waste dried by oven and lyophilizer), and the Fourier-transform infrared spectroscopy profile of the chitin obtained from the deproteinization process displayed the characteristic of chitin. Finally, the obtained chitin exhibited an effect comparable to commercial chitin in terms of adsorption against Congo Red (90.48% and 90.91%, respectively). Thus, P. elgii TKU051 showed potential in the reclamation of chitinous fishery by-products for proteases production and chitin extraction.
Assuntos
Quitina/química , Pesqueiros , Paenibacillus/metabolismo , Peptídeo Hidrolases/biossíntese , Resíduos , Adsorção , Animais , Compostos de Bifenilo/química , Corantes/química , Vermelho Congo/química , Decápodes , Decapodiformes , Fermentação , Picratos/química , Reciclagem , Poluentes Químicos da Água , alfa-Glucosidases/metabolismoRESUMO
The chitinase producing strain TKU008 was isolated from the soil in Taiwan, and it was identified as a new species of Pseudomonas. The culture condition suitable for production of chitinase was found to be shaken at 30 degrees C for 4 days in 100 mL of medium containing 1% shrimp and crab shell powder, 0.1 % K2HPO4 and 0.05% MgSO4 . 7 H2O (pH 7). The TKU008 chitinase was suppressed by the simultaneously existing protease, which also showed the maximum activity at the forth day of incubation. The molecular mass of the chitinase was estimated to be 40 kDa by SDS-PAGE. The optimum pH, optimum temperature, pH stability, and thermal stability of the chitinase were pH 7, 50 degrees C, pH 6-7, and 50 degrees C, respectively. The chitinase was completely inhibited by Mn2+ and Cu2+. The results of peptide mass mapping showed that eleven tryptic peptides of the chitinase were identical to a chitinase CW from Bacillus cereus (GenBank accession number gi 45827175) with a 32 % sequence coverage.
Assuntos
Proteínas de Bactérias/química , Proteínas de Bactérias/isolamento & purificação , Quitinases/química , Quitinases/isolamento & purificação , Pseudomonas/enzimologia , Sequência de Aminoácidos , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Quitinases/genética , Quitinases/metabolismo , Estabilidade Enzimática , Dados de Sequência Molecular , Peso Molecular , Pseudomonas/química , Pseudomonas/genética , Pseudomonas/isolamento & purificação , Microbiologia do SoloRESUMO
As a green and effective technique in the production of a large number of valuable products, the microbial conversion of chitinous fishery wastes is receiving much attention. In this study, protease production using the Paenibacillus mucilaginosus TKU032 strain was conducted on culture media containing several common types of chitinous fishery by-products serving as the carbon and nitrogen (C/N) nutrition source. Among the chitinous wastes, 1.5% (w/v) shrimp head powder (SHP) was found to be the most appropriate nutritional source for protease production when a maximal enzyme activity of 3.14 ± 0.1 U/mL was observed on the 3rd day of the culture period. The molecular mass of P. mucilaginosus TKU032 protease was estimated to be nearly 32 kDa by the polyacrylamide gel electrophoresis method. The residual SHP obtained from the culture medium was also considered to be utilized for chitin extraction. The deproteinization rate of the fermentation was estimated to be 45%, and the chitin obtained from fermented SHP (fSHP) displayed a similar characteristic Fourier-transform infrared spectroscopy (FTIR) profile as that from SHP. In addition, SHP, fSHP, and chitins obtained from SHP and fSHP were investigated for their adsorptive capacity of nine types of dyes, and chitin obtained from fSHP displayed a good adsorption rate on Congo Red and Red No. 7, at 99% and 97%, respectively. In short, the results provide potential support for the utilization of SHP in the production of P. mucilaginosus TKU032 protease via the fermentation as well as the preparation of chitin from fSHP as an effective dye adsorbent.
RESUMO
A chitinase (CHT1) and a chitosanase (CHS1) were purified from the culture supernatant of Pseudomonas sp. TKU015 with shrimp shell wastes as the sole carbon and nitrogen source. The optimized conditions of this new species strain (Gen Bank Accession Number EU103629) for the production of chitinases were found to be when the culture was shaken at 30 degrees C for 3 days in 100 mL of medium (pH 8) containing 0.5% shrimp shell powder (SSP) (w/v), 0.1% K2HPO4, and 0.05% MgSO(4).7H2O. The molecular weights of CHT1 and CHS1 determined by SDS-PAGE were approximately 68 kDa and 30 kDa, respectively. The optimum pH, optimum temperature, pH stability, and the thermal stability of CHT1 and CHS1 were pH 6, 50 degrees C, pH 5-7, <50 degrees C and pH 4, 50 degrees C, pH 3-9, <50 degrees C, respectively. CHT1 was inhibited completely by Mn2+ and Fe2+, and CHS1 was inhibited by Mn2+, Cu2+, and PMSF. CHT1 was only specific to chitin substrates, whereas the relative activity of CHS1 increased when the degree of deacetylation of soluble chitosan increased.
Assuntos
Carbono/metabolismo , Quitinases/isolamento & purificação , Decápodes , Glicosídeo Hidrolases/isolamento & purificação , Nitrogênio/metabolismo , Pseudomonas/enzimologia , Acetilação , Animais , Quitina/farmacologia , Quitinases/química , Quitinases/metabolismo , Quitosana/metabolismo , Meios de Cultura , Compostos Ferrosos/farmacologia , Glicosídeo Hidrolases/química , Glicosídeo Hidrolases/metabolismo , Concentração de Íons de Hidrogênio , Compostos de Manganês/farmacologia , Dados de Sequência Molecular , Peso Molecular , Sefarose/análogos & derivados , Sefarose/química , Especificidade por Substrato , TemperaturaRESUMO
A protease-producing bacterium was isolated and identified as Chryseobacterium indologenes TKU014. The optimized condition for protease production was found when the culture was shaken at 30 degrees C for one day in 50 mL of medium containing 0.5% shrimp shell powder (w/v), 0.1% K(2)HPO(4), and 0.05% MgSO(4).7H(2)O. Three extracellular proteases (P1, P2, and P3) were purified from culture by DEAE-Sepharose and Phenyl Sepharose chromatography. Three enzymes all showed activities of keratinase and elastase with molecular weights of 56, 40, 40 kDa, respectively. The inhibitory effect of metal chelator EDTA and Zn-specific chelator 1,10-phenanthroline characterized three enzymes as Zn-metalloproteases. Peptide mass fingerprints of P1, P2, and P3 were determined by using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Similarity search in the NCBI non-redundant protein sequence database revealed that three enzymes exhibited no significant homology to any other reported microbial peptides. Therefore, P1, P2, and P3 are most likely novel proteins.
Assuntos
Anostraca/microbiologia , Chryseobacterium/enzimologia , Metaloproteases/isolamento & purificação , Metaloproteases/metabolismo , Animais , Proteínas de Bactérias/isolamento & purificação , Proteínas de Bactérias/metabolismo , Cromatografia por Troca Iônica , Chryseobacterium/isolamento & purificação , Meios de Cultura , Cinética , Frutos do Mar/microbiologia , Microbiologia do SoloRESUMO
Paenibacillus macerans TKU029 can produce exopolysaccharides (EPSs; 3.46 g/L) and a biosurfactant (1.78 g/L) in a medium with 2 % (w/v) squid pen powder as the sole carbon/nitrogen source. The biosurfactant can reduce the surface tension of water from 72.30 to 35.34 mN/m at a concentration of 2.76 g/L and reach an emulsification index of 56 % after a 24-h reaction with machine oil. This biosurfactant is stable at 121 °C for 20 min, over a pH range from 3 to 11, and in <5 % salt solutions. It also shows significant antimicrobial activity, which remains active after treatment at 121 °C and at pH values from 4 to 10, against Escherichia coli BCRC13086, Staphylococcus aureus BCRC10780, Fusarium oxysporum BCRC32121 and Aspergillus fumigatus BCRC30099. Furthermore, human skin shows from 37.3 to 44.3 % hydration after being treated with TKU029 EPSs for 180 min. These results imply that EPSs and the biosurfactant from this strain have potential in cosmetics, for removal of oil contamination, and as antimicrobial agents.
Assuntos
Anti-Infecciosos/farmacologia , Paenibacillus/metabolismo , Polissacarídeos Bacterianos/biossíntese , Polissacarídeos Bacterianos/farmacologia , Tensoativos/farmacologia , Bactérias/efeitos dos fármacos , Emulsões/química , Feminino , Fungos/efeitos dos fármacos , Humanos , Concentração de Íons de Hidrogênio/efeitos dos fármacos , Micelas , Testes de Sensibilidade Microbiana , Paenibacillus/efeitos dos fármacos , Paenibacillus/crescimento & desenvolvimento , Salinidade , Tensão Superficial/efeitos dos fármacos , Temperatura , Fatores de Tempo , Adulto JovemRESUMO
Using squid pen powder (SPP) as the sole C/N source, Paenibacillus sp. TKU023 produced exopolysaccharides (EPS) and antioxidant. With medium containing 1.5% SPP, 0.1% K(2)HPO(4), and 0.05% MgSO(4)·7H(2)O, pH 7.23, the culture was incubated at 37°C in liquid (50 mL/250 mL) for five days. The resultant culture supernatant had higher EPS productivity (4.55 g/L). The crude EPS were isolated by centrifugation, methanol precipitation and deproteinization. The characterization of the EPS demonstrated that it was mainly composed of glucose and maltose. In addition, the culture supernatant incubated for four days by using baffled base flask showed the strongest antioxidant activities and the highest total phenolic content, but maximum EPS production was found at the fifth day by using flat base flask. The production of two invaluable environmental-friendly biomaterials (EPS and antioxidant) is unprecedented. Besides, the use of SPP (waste) is green that made the whole process more valuable and attractive.