Partial Characterization of an Acidic Protease from Rhizopus stolonifer RN-11

Liu, Na; Huang, Liang

doi:10.2174/1874070701509010199

Cited by 4 publications

(2 citation statements)

References 20 publications

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“…respectively. Griffen et al [29] also reported that aspartic protease by F. graminearum had molecular mass of 40 kDa.With respect to results obtained, a similar protein size of 70 kDa on SDS-PAGE was reported by Na and Liang [30].…”

Section: Application Of Proteasesupporting

confidence: 88%

Purification, Characterization and De-Staining Potentials of a Thermotolerant Protease Produced by Fusarium oxysporum

Ja’afaru

Chimbekujwo

Ajunwa

2019

Period. Polytech. Chem. Eng.

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Proteases are important industrial enzymes and fungi prove to be good sources of such enzymes. Purification techniques are however necessary for increased specificity in activity and better industrial value. Based on this, a protease produced by a Fusarium oxysporum was purified to homogeneity by Sephadex G-200 column and α-casein agarose chromatography. The enzyme had a molecular weight of 70 kDa in SDS-PAGE. Purified Fusarium oxysporum protease had a specific activity of 93.88 U/mg protein. The purification magnitude was 7.7 and the total yield was 20 %. Purified protease had an optimum pH of 5.0 while the optimum temperature was 40 °C.The enzyme was also thermotolerant (approximately 100 % at 40 °C for 2 h). The enzyme activity was stimulated by surfactants and metal ions like, Tween-20 and Mg 2+ . Enzyme activity was inhibited in presence of PMSF and EDTA. Casein was found to be the best substrate for protease activity of Fusarium oxysporum FWT1. Protease were tested upon blood stain for de-clotting of blood and was found to exhibit good de-clotting and de-staining activity after 15 minutes treatment time. KeywordsFusarium oxysporum, protease, enzyme purification, thermotolerant 2| Ja'afaru et al.Period. Polytech. Chem. Eng.

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Section: Application Of Proteasesupporting

confidence: 88%

Purification, Characterization and De-Staining Potentials of a Thermotolerant Protease Produced by Fusarium oxysporum

Ja’afaru

Chimbekujwo

Ajunwa

2019

Period. Polytech. Chem. Eng.

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“…On the other hand, NiCl 2 and ZnSO 4 showed an inhibitory effect on the milkclotting activity as compared to the control (Skim-milk only). Similarly, Ca 2+ (El-Tanboly et al, 2013), Na + , K + , Mn 2+ and Ca 2+ (Liu and Huang, 2015) and Ca 2+ , Mg 2+ , Mn 2+ (Kumari et al 2016) and Mn 2+ , Ca 2 (Ahmed et al, 2016) were shown a noticeable effect on enhancing the the activity of milk-clotting enzyme from M. pusillus QM 436, acidic protease from Rhizopus stolonifer, MCE from B. subtilis MTCC 10422 and MCE from B. stearothermophilus, respectively. Whereas the addition of Co 2+ , Cd 2+ and Zn 2+ were found to have an inhibitory effect on the activity of MCE from B. subtilis MTCC 10422 (Kumari et al 2016).…”

Section: Discussionmentioning

confidence: 92%

Purification and Characterization of Aspartic Protease Produced fromAspergillus oryzaeDRDFS13 under Solid-State Fermentation

Mamo

Orellana

Yelemane

et al. 2020

Preprint

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Aspartic proteases (E.C.3.4.23.) are endopeptidases with molecular masses ranging between 30-45 kDa. They depend on aspartic acid residues for their catalytic activity and show maximal activity at low pH. Thus the main objective of the present study was to purify and characterize aspartic protease from locally identified fungi by solid-state fermentation. The aspartic protease in the current study was obtained from A. oryzae DRDFS13 under SSF. The crude enzyme extract was purified by size-exclusion (SEC) and ion-exchange (IEC) chromatography. The protein contents of crude enzyme and IEC fractions were determined by BCA methods while the presence of N-glycosylation was checked using Endo-H. Inhibition studies were conducted using protease inhibitors. The milk-clotting activity (MCA), protease activity (PA); molecular weight and enzyme kinetics were determined using standard methods. Optimum temperature and stability, optimum pH and stability, and the effect of cations on MCA were assessed using standard methods. The maximum MCA (477.11 U/mL) was recorded from IEC fraction A8. The highest specific activity (183.50 U/mg), purification fold (6.20) and yield (9.2%) were also obtained from the same fraction (IEC A8). The molecular weight of 40 kDa was assigned for the purified enzyme (IEC A8). However, its molecular weight was decreased to 30 KDa upon deglycosylation assay which infers that the protein is glycosylated. Incubation of the pure enzyme (IEC A8) with pepstatin A caused a 94 % inhibition on MCA. The dialyzed enzyme showed a Km and Vmax values of 17.50 mM and 1369 U, respectively. The enzyme showed maximum MCA at 60 oC and pH 5.0 with stability at pH 4.5-6.5 and temperature 35-45 oC. Most cat-ions stimulate the activity of the enzyme; moreover, the highest MCA was detected at 50 mM of MnSO4. Furthermore, the results obtained in the present study confirmed that the aspartic protease enzyme produced from A. oryzae DRDFS13 and purified in ion-exchange chromatography could be used as a substitute source of rennet enzyme for cheese production.

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Purification and Characterization of Aspartic Protease Produced from Aspergillus oryzae DRDFS13 MN726447 under Solid-State Fermentation

et al. 2021

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Partial Characterization of an Acidic Protease from Rhizopus stolonifer RN-11

Cited by 4 publications

References 20 publications

Purification, Characterization and De-Staining Potentials of a Thermotolerant Protease Produced by Fusarium oxysporum

Purification, Characterization and De-Staining Potentials of a Thermotolerant Protease Produced by Fusarium oxysporum

Purification and Characterization of Aspartic Protease Produced fromAspergillus oryzaeDRDFS13 under Solid-State Fermentation

Purification and Characterization of Aspartic Protease Produced from Aspergillus oryzae DRDFS13 MN726447 under Solid-State Fermentation

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