Proteolytic and Lipolytic Activity of Molds Isolated from Aged Beef

1982 ◽  
Vol 45 (13) ◽  
pp. 1242-1244 ◽  
Author(s):  
A. W. KOTULA ◽  
S. G. CAMPANO ◽  
D. M. KINSMAN
Keyword(s):  
Proteolytic Activity ◽  
Colony Size ◽  
Lipolytic Activity ◽  
Skim Milk ◽  
Tween 80 ◽  
Size Ratio ◽  
Skim Milk Agar ◽  
Mucor Mucedo

This study evaluated the proteolytic and lipolytic activity of several strains of Thamnidium elegans, Mucor mucedo and Chaetostylum fresenii on selected test proteins and lipids. At 18°C, the zone of hydrolysis to colony size ratio on skim milk agar, representing proteolytic activity after 4 d, was 0.92, 0.80 and 0.67 for M. mucedo, C. fresenii and T. elegans, respectively. A similar trend was noted after 4 d of incubation at 24°C. There was positive lipolytic activity on Tween 80 at 18 and 24°C for the same three molds. The proteolytic and lipolytic activity decreased with decreasing temperatures so that at 4°C, the temperature of most probable use if applied to meat, the effect was negligible unless long incubation times were used. The absence of proteolytic activity of the molds at 4°C and the impracticality of aging beef at 18 or 24°C suggest that treatment of meat with molds to enhance tenderness may not be feasible.

scholarly journals Complex regulation of AprA metalloprotease in Pseudomonas fluorescens M114: evidence for the involvement of iron, the ECF sigma factor, PbrA and pseudobactin M114 siderophore

Microbiology ◽  
2006 ◽  
Vol 152 (1) ◽  
pp. 29-42 ◽  
Author(s):  
Bláithín Maunsell ◽  
Claire Adams ◽  
Fergal O'Gara
Keyword(s):  
Pseudomonas Fluorescens ◽  
Proteolytic Activity ◽  
Yeast Extract ◽  
Sigma Factor ◽  
Skim Milk ◽  
Protease Production ◽  
Iron Starvation ◽  
Transcription Analysis ◽  
Skim Milk Agar ◽  
Complex Regulation

In the soil bacterium Pseudomonas fluorescens M114, extracellular proteolytic activity and fluorescent siderophore (pseudobactin M114) production were previously shown to be co-ordinately negatively regulated in response to environmental iron levels. An iron-starvation extracytoplasmic function sigma factor, PbrA, required for the transcription of siderophore biosynthetic genes, was also implicated in M114 protease regulation. The current study centred on the characterization and genetic regulation of the gene(s) responsible for protease production in M114. A serralysin-type metalloprotease gene, aprA, was identified and found to encode the major, if not only, extracellular protease produced by this strain. The expression of aprA and its protein product were found to be subject to complex regulation. Transcription analysis confirmed that PbrA was required for full aprA transcription under low iron conditions, while the ferric uptake regulator, Fur, was implicated in aprA repression under high iron conditions. Interestingly, the iron regulation of AprA was dependent on culture conditions, with PbrA-independent AprA-mediated proteolytic activity observed on skim milk agar supplemented with yeast extract, when supplied with iron or purified pseudobactin M114. These effects were not observed on skim milk agar without yeast extract. PbrA-independent aprA expression was also observed from a truncated transcriptional fusion when grown in sucrose asparagine tryptone broth supplied with iron or purified pseudobactin M114. Thus, experimental evidence suggested that iron mediated its effects via transcriptional activation by PbrA under low iron conditions, while an as-yet-unidentified sigma factor(s) may be required for the PbrA-independent aprA expression and AprA proteolytic activity induced by siderophore and iron.

Proteolysis byLactobacillus fermentumIFO3956 isolated from Egyptian milk products decreases immuno-reactivity of αS1-casein

2011 ◽  
Vol 78 (2) ◽  
pp. 203-210 ◽  
Author(s):  
Shady El-Ghaish ◽  
Hanitra Rabesona ◽  
Yvan Choiset ◽  
Mahmoud Sitohy ◽  
Thomas Haertlé ◽  
...  
Keyword(s):  
Proteolytic Activity ◽  
Skim Milk ◽  
Tween 80 ◽  
Molecular Forms ◽  
Allergic Reactions ◽  
Terminal Portion ◽  
Phosphorylated Peptides ◽  
Casein Hydrolysates ◽  
Free Media ◽  
Hydrolysis Of

Proteinase activity ofLactobacillus fermentumIFO3956 cells was higher when they were grown on milk-based media than on 10% reconstituted skim milk. The lowest protease activity was observed when cells were grown on milk-free media. The extraction of milk-induced cell-bound proteases fromLb. fermentumIFO3956 was most efficient using 1% Tween 80 while the use of 1% SDS inhibited all proteolytic activity. Two bands of ∼35 and >100 kDa were observed by zymogram, indicating that proteolytic activity corresponded to the presence of at least two types of enzymes or two molecular forms of one enzyme. Mass spectrometry analyses of αS1-casein hydrolysates detected 24 peptides with sizes ranging from 5 to 36 amino acids, including 9 phosphorylated peptides, resulting from the fermentation ofLb. fermentumIFO3956 of αS1-casein. Most of the identified peptides originated from the N-terminal portion of αS1-casein. The studied bacterial strain could hydrolyze αS1-casein in many sites including the epitopes triggering the allergic reactions against αS1-caseine.g.at the positions 23, 30, 41, 71, 91, 98, 126, 179. After hydrolysis of αS1-casein withLb. fermentumIFO3956 the recognition and the binding of this casein to IgE from the pooled sera of 18 patients with cow's milk allergy was significantly reduced.

Studies on the Production of Acid Protease by Submerged Fermentation

2009 ◽  
Vol 5 (1) ◽  
Author(s):  
Shishir Sinha ◽  
Shalini Sinha
Keyword(s):  
Enzyme Activity ◽  
Proteolytic Activity ◽  
Enzyme Production ◽  
Milk Powder ◽  
Skim Milk ◽  
Peanut Meal ◽  
Tween 80 ◽  
Skim Milk Powder ◽  
Acid Protease ◽  
Aspergillus Awamori

Proteases are enzymes that hydrolyze peptide bonds and, therefore, lead to the disassembly of proteins. Commercially these are extremely important as more than 60% of the total enzyme market is made up of proteases, out of which 40% are acid proteases. The objective of this study was to compare the available standard strains, Rhizopus oligosporus MTCC-556, Rhizomucor miehei MTCC-546 and Aspergillus awamori MTCC-548, which were examined for the production of acid protease by submerged fermentation. Aspergillus awamori showed maximum proteolytic activity and was selected for further optimization studies. During the course of study the medium was altered. Effect of different carbon sources (lactose, sucrose, and combination of these two in same ratios and glucose) on the proteolytic activity of acid protease produced by A. awamori MTCC 548 was studied and it was found that glucose showed highest proteolytic activity. The effect of various concentrations of glucose was also studied on the acid protease production and its 1% concentration was found to be optimum; it showed proteolytic activity of 0.11 U/ml. Among the different nitrogen sources, such as casein, peptone, skim-milk powder and peanut meal, the peanut meal was found better for enzyme production. Peanut meal, with a concentration of .2% in the medium, increased proteolytic activity up to 0.218 U/ml. The effect of additives, such as Tween-80 and chemicals like CaCl2 and skim-milk powder, was also studied and it was found that 0.05% Tween-80 was effective in enzyme production and a proteolytic activity of 0.225 U/ml was obtained. The enzyme extract was separated in the form of supernatant by using centrifuge and the enzyme activity was analyzed by Anson’s method using a spectrophotometer. The enzyme produced was recovered by using a saturated solution of 80% ammonium sulphate and protein content was determined by Lowery method using a spectrophotometer. It was found that the specific enzyme activity was increased from 0.155 U/mg proteins to 0.174 U/mg proteins showing a purification fold by a factor of 1.12 by using 80% saturated ammonium sulphate.

scholarly journals Chromatographic Characterization and In Vitro Bioactivity Evaluation of Lactobacillus helveticus Hydrolysates upon Fermentation of Different Substrates

2021 ◽  
Vol 11 (2) ◽  
pp. 811
Author(s):  
Federica Ianni ◽  
Alessandra Anna Altomare ◽  
Beniamino T. Cenci-Goga ◽  
Francesca Blasi ◽  
Luca Grispoldi ◽  
...  
Keyword(s):  
Liquid Chromatography ◽  
Proteolytic Activity ◽  
Bioactive Peptides ◽  
Biological Activities ◽  
Skim Milk ◽  
Brain Heart Infusion ◽  
Starter Cultures ◽  
Lactobacillus Helveticus ◽  
Food Sources

Among various food sources, milk proteins remain the major vector for functional peptides endowed with several biological activities. Particularly, the proteolytic activity of lactic acid bacteria during milk fermentation has been one of the most followed strategies to produce bioactive peptides. In the present study, the exploration of the activity of several starter cultures, at different fermentation times, was firstly investigated by reversed phase-high performance liquid chromatography. Among the tested strains, Lactobacillus helveticus showed a higher proteolytic activity and it was submitted to further investigations by changing the fermentation substrate (skim milk, brain heart infusion, peptone water) as well as the extraction strategy (trichloroacetic acid vs. glass beads). The chromatographic analyses and the in vitro antioxidant and antihypertensive assays highlighted considerable differences for L. helveticus hydrolysates from different substrates, while a negligible impact by the two extraction protocols emerged. Furthermore, nano-high pressure liquid chromatography coupled with a high resolution mass spectrometry analyzer allowed the preliminary discrimination of fractions from fermented skim milk, likely responsible for the found activity. The obtained results suggest the possibility of varying the fermentation parameters in order to maximize the functional effects of the bioactive peptides.

LIPOLYTIC AND PROTEOLYTIC ACTIVITY OF ENTEROCOCCI AND LACTIC GROUP STREPTOCOCCI ISOLATED FROM YOUNG CHEDDAR CHEESE1

1970 ◽  
Vol 33 (9) ◽  
pp. 365-372 ◽  
Author(s):  
A. M. Dovat ◽  
G. W. Reinbold ◽  
E. G. Hammond ◽  
E. R. Vedamuthu
Keyword(s):  
Acetic Acid ◽  
Proteolytic Activity ◽  
Lipolytic Activity ◽  
Cheddar Cheese ◽  
Streptococcus Faecalis ◽  
Screening Techniques ◽  
Ph Values ◽  
Plate Counts

Lipolytic and proteolytic screening techniques were applied to cultures isolated from young Cheddar cheese manufactured in 10 Iowa cheese plants. Twenty-one cultures were selected for study. These included 16 enterococci and 5 lactic group streptococci. These strains were examined for lipolytic activity when grown in skimmilk, cream, and skimmilk containing tributyrin; changes in proteolysis index, plate counts, and pH in skimmilk incubated at 7, 15, 21, and 32 C also were determined, And, combinations of enterococci and lactic streptococci were studied. One-half of the Streptococcus durans strains frequently produced as much as 10 times more acetic acid than the others; the five strains of lactic streptococci consistently produced the lowest quantities of acetic acid. Compared with enterococci, except for Streptococcus faecalis var. liquefaciens, the lactic streptococci were more proteolytic, produced lower pH values, and had less viability at 15, 21, and 32 C. Enterococci other than S. faecalis var. liquefaciens were not proteolytic. All cultures showed tributyrinase activity; enterococci were the most active. Combining enterococci and lactic streptococci produced anomalous results.

Proteolytic and Lipolytic Activities of some Toxigenic and Nontoxigenic Aspergilli and Penicillia

1980 ◽  
Vol 43 (5) ◽  
pp. 354-355 ◽  
Author(s):  
S. M. EL-GENDY ◽  
E. H. MARTH
Keyword(s):  
Proteolytic Activity ◽  
Aspergillus Flavus ◽  
Aspergillus Parasiticus ◽  
Lipolytic Activity ◽  
The Other ◽  
Aspergillus Ochraceus ◽  
Penicillium Roqueforti ◽  
Two Cultures ◽  
Penicillium Cyclopium ◽  
Penicillium Patulum

Eighteen strains of Aspergillus flavus or Aspergillus parasiticus, one of Aspergillus ochraceus and 12 strains or species of Penicillium, many of them isolated from cheese, were evaluated for their proteolytic and lipolytic activities. Strains of A. flavus exhibited considerable proteolytic and little lipolytic activity, whereas the reverse was true for strains of A. parasiticus. Of the Penicillium cultures tested, 10 exhibited considerable lipolytic activity, but only five had marked proteolytic activity. Two cultures, Penicillium patulum M59, and Penicillium cyclopium No. 8, were markedly lipolytic and proteolytic. Of the other cultures, greatest lipolytic activity was associated with Penicillium roqueforti 849, Penicillium puberulum No. 33, A. parasiticus NRRL 3145 and NRRL 465 and A. ochraceus NRRL 3174, whereas greatest proteolytic activity of all the cultures was associated with P. patulum M59, P. cyclopium No. 25 and A. flavus WB500, 4018, 4098 and NRRL 5565.

Lipolytic Activity During Storage of Human Milk: Accumulation of Free Fatty Acids1

1984 ◽  
Vol 47 (9) ◽  
pp. 690-693 ◽  
Author(s):  
C. W. DILL ◽  
C. T. CHEN ◽  
E. S. ALFORD ◽  
R. L. EDWARDS ◽  
R. L. RICHTER ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Room Temperature ◽  
Lipolytic Activity ◽  
Skim Milk ◽  
Milk Samples ◽  
Fatty Acid Accumulation ◽  
Whole Milk ◽  
Freeze Dried ◽  
Rapid Accumulation ◽  
Acid Accumulation

Lipolysis was quantitated during storage of fluid and freezedried human whole and skim milks. Fatty acid accumulation was faster in whole fluid milk stored for 1 week at 4°C than in frozen (−20°C) samples stored for 180 d. The rapid accumulation of fatty acids during 24 h of storage at 4°C was enhanced in previously frozen milk samples. While freeze-dried whole milk showed no lipolysis when stored at −20°C, accumulation of free fatty acids was rapid in samples stored at room temperature. Fluid and freeze-dried skim milk samples exhibited no appreciable lipolysis.

scholarly journals Shelf life extension of liquid whole eggs by heat and bacteriocin treatment

2010 ◽  
Vol 28 (No. 4) ◽  
pp. 280-289 ◽  
Author(s):  
P. Miller ◽  
M.E. Haveroen ◽  
K. Solichová ◽  
R. Merkl ◽  
L.M. McMullen ◽  
...  
Keyword(s):  
Lactic Acid ◽  
Shelf Life ◽  
Proteolytic Activity ◽  
Inhibitory Concentration ◽  
Lipolytic Activity ◽  
Life Extension ◽  
High Proteolytic Activity ◽  
Liquid Whole Egg ◽  
Spoilage Microflora ◽  
Whole Egg

During a 15-month period, samples of commercially pasteurised liquid whole egg (LWE) were tested for the presence of spoilage microflora. The total bacterial counts were 2.2 ± 0.6 log CFU/g and total lactic acid bacteria (LAB) counts were 1.9 ± 0.6 log CFU/g. Enterobacteriaceae were detected in 2 samples. Out of the tested samples, 45 LAB were isolated and identified, with 30 strains identified as Enterococcus faecium, 12 as Enterococcus faecalis, and 3 as Lactobacillus paracasei subsp. paracasei. All strains, except 6 strains of E. faecium, possessed lipolytic activity. All the E. faecalis strains and one strain of E. faecium showed a high proteolytic activity, while moderate proteolytic activity was shown by 3 lactobacilli strains. Minimum inhibitory concentration (MIC) of nisin and Micocin X was measured against groups of isolated strains, and ranged from 10.4 µg/ml to 41.7 µg/ml for nisin and from 0.2 mg/ml to 1.6 mg/ml for Micocin X. The LWEs supplemented with 6.25 mg/l of nisin or with 500 mg/ml of Micocin X were pasteurised at 65°C for 2.5 minutes. The shelf life of LWE with the addition of nisin or Micocin X stored under refrigerator conditions was extended by a minimum of 5 weeks.

scholarly journals PRODUCTION OF KERATINASES FROM NOCARDIOPSIS SP. 28ROR AS A NOVEL IRAQI STRAIN

2017 ◽  
Vol 10 (4) ◽  
pp. 160
Author(s):  
Rabab Omran Al-jelawi-
Keyword(s):  
Potential Application ◽  
Skim Milk ◽  
Gene Analysis ◽  
Poultry Farm ◽  
Chicken Feather ◽  
Feather Meal ◽  
Skim Milk Agar ◽  
16S R Rna ◽  
Acid Type

Objectives: isolate a novel feather- degrading actinobacterial species had the ability to produce wide pH activity keratinases.Methods: Of 23 actinobacterial isolates were recovered from farm soil, poultry farm soil and feather wastes, these isolates were screened for protease and keratinase production on skim milk agar, feather  media, and  antimicrobial production. One potential  isolate was identified depending on phenotypical, physiological and molecular according to partial sequences of 16S r RNA gene analysis and optimized  keratinase production. Results:   11 isolates out of 22 protease producer  had the ability to degrade raw chicken feather and some of these  isolates produced  antifungal and antibacterial metabolites.The potential isolate,  Nocardiopsis sp. 28ROR (GenBank: KC702802.1), produced two types of extracellular keratinases in feather meal  medium at pH6 (acid type), 30-35°C  for 7d  and  the alkaline keratinase at pH10, 40°C  for 7d.Conclusion: The Nocardiopsis sp. 28ROR was a novel strain produced keratinases using feather meal degradation as a cheap waste medium. The wide tolerance of temperature and pH by keratinase makes it an ideal contender to be investigated further for potential application as a detergent additive.Keywords: Nocardiopsis, Keratinase, Optimization, Feather medium, Antibiotic. 

scholarly journals Isolation and Characterization of Metalloproteases with a Novel Domain Structure by Construction and Screening of Metagenomic Libraries

2009 ◽  
Vol 75 (8) ◽  
pp. 2506-2516 ◽  
Author(s):  
Tanja Waschkowitz ◽  
Stephanie Rockstroh ◽  
Rolf Daniel
Keyword(s):  
Proteolytic Activity ◽  
Signal Sequence ◽  
Skim Milk ◽  
Environmental Dna ◽  
Average Insert Size ◽  
Extracellular Proteases ◽  
Insert Size ◽  
T4 Dna Ligase ◽  
Metagenomic Libraries ◽  
Isolation And Characterization

ABSTRACT Small-insert metagenomic libraries from four samples were constructed by a topoisomerase-based and a T4 DNA ligase-based approach. Direct comparison of both approaches revealed that application of the topoisomerase-based method resulted in a higher number of insert-containing clones per μg of environmental DNA used for cloning and a larger average insert size. Subsequently, the constructed libraries were partially screened for the presence of genes conferring proteolytic activity. The function-driven screen was based on the ability of the library-containing Escherichia coli clones to form halos on skim milk-containing agar plates. The screening of 80,000 E. coli clones yielded four positive clones. Two of the plasmids (pTW2 and pTW3) recovered from positive clones conferred strong proteolytic activity and were studied further. Analysis of the entire insert sequences of pTW2 (28,113 bp) and pTW3 (19,956 bp) suggested that the DNA fragments were derived from members of the genus Xanthomonas. Each of the plasmids harbored one gene (2,589 bp) encoding a metalloprotease (mprA, pTW2; mprB, pTW3). Sequence and biochemical analyses revealed that MprA and MprB are similar extracellular proteases belonging to the M4 family of metallopeptidases (thermolysin-like family). Both enzymes possessed a unique modular structure and consisted of four regions: the signal sequence, the N-terminal proregion, the protease region, and the C-terminal extension. The architecture of the latter region, which was characterized by the presence of two prepeptidase C-terminal domains and one proprotein convertase P domain, is novel for bacterial metalloproteases. Studies with derivatives of MprA and MprB revealed that the C-terminal extension is not essential for protease activity. The optimum pH and temperature of both proteases were 8.0 and 65°C, respectively, when casein was used as substrate.

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