Pharmacological potential of plant proteases inhibitors

Las proteasas constituyen un conjunto de enzimas que juegan un papel crucial en numerosos procesos celulares, tales como la degradación de proteínas implicadas en la regulación del ciclo celular, la apoptosis, el crecimiento, la activación, adhesión, invasión y migración celular, las metástasis, las interacciones celulares, la transducción de señales, la fagocitosis y la angiogénesis, entre otros. A nivel tisular la relación entre proteasas y sus inhibidores mantiene un equilibrado balance, equilibrio que se quiebra en casos de carcinogénesis, infecciones y otros desórdenes patológicos. En efecto, las proteasas juegan un papel clave en la patogénesis. Un gran número de desórdenes fisiopatológicos resultan del desbalance de la actividad proteolítica. En este marco los inhibidores de proteasas (IPs) son parte elemental del sistema de defensa endógeno, ya que ayudan a regular y balancear las actividades proteolíticas celulares. Se ha demostrado que los IPs evitan la progresión de tumores y metástasis y también tienen el potencial de contrarrestar desórdenes hereditarios tales como el enfisema y la epilepsia. Los IPs también pueden interferir el ciclo de vida de muchos virus y en consecuencia ayudar a prevenir algunas patologías virales. Por su parte los IPs sintéticos son integrantes de la terapia combinada contra el SIDA y tienen un enorme potencial para ser utilizados contra muchas otras enfermedades. De allí que los IPs, tanto naturales como sintéticos, resultan una alternativa válida para enriquecer el arsenal farmacológico existente a la fecha.Proteases represent an enzyme group that plays a crucial role in many cell processes, such as degradation of proteins involved in the regulation of cell cycle, apoptosis, cell growth and activation; adhesion, invasion, and cell migration; metastasis, cellular interactions, signal transduction, phagocytosis, and angiogenesis, among others. At a tisular level, the relationship between proteases and their inhibitors maintains an adjusted balance, which is broken in cases of carcinogenesis, infections and other pathological disorders. Indeed, proteases play a key role in pathogenesis. A large number of physiopathological disorders result from the imbalance of proteolytic activity. In this context protease inhibitors (PIs) are a key component of the endogenous defense system, helping to regulate and balance the cellular proteolytic activities. It has been shown that PIs prevent tumor progression and metastasis and also have the potential to counteract hereditary disorders such as emphysema and epilepsy. PIs may also interfere with the life cycle of many viruses and thus help to prevent some viral diseases. Synthetic PIs are part of a combined therapy against AIDS and have enormous potential for being used against many other diseases. Hence PIs, both natural and synthetic, are a valid way to enrich the pharmacological arsenal existing at the date.Facultad de Ciencias Exacta

Pharmacological potential of plant proteases inhibitors

Las proteasas constituyen un conjunto de enzimas que juegan un papel crucial en numerosos procesos celulares, tales como la degradación de proteínas implicadas en la regulación del ciclo celular, la apoptosis, el crecimiento, la activación, adhesión, invasión y migración celular, las metástasis, las interacciones celulares, la transducción de señales, la fagocitosis y la angiogénesis, entre otros. A nivel tisular la relación entre proteasas y sus inhibidores mantiene un equilibrado balance, equilibrio que se quiebra en casos de carcinogénesis, infecciones y otros desórdenes patológicos. En efecto, las proteasas juegan un papel clave en la patogénesis. Un gran número de desórdenes fisiopatológicos resultan del desbalance de la actividad proteolítica. En este marco los inhibidores de proteasas (IPs) son parte elemental del sistema de defensa endógeno, ya que ayudan a regular y balancear las actividades proteolíticas celulares. Se ha demostrado que los IPs evitan la progresión de tumores y metástasis y también tienen el potencial de contrarrestar desórdenes hereditarios tales como el enfisema y la epilepsia. Los IPs también pueden interferir el ciclo de vida de muchos virus y en consecuencia ayudar a prevenir algunas patologías virales. Por su parte los IPs sintéticos son integrantes de la terapia combinada contra el SIDA y tienen un enorme potencial para ser utilizados contra muchas otras enfermedades. De allí que los IPs, tanto naturales como sintéticos, resultan una alternativa válida para enriquecer el arsenal farmacológico existente a la fecha.Proteases represent an enzyme group that plays a crucial role in many cell processes, such as degradation of proteins involved in the regulation of cell cycle, apoptosis, cell growth and activation; adhesion, invasion, and cell migration; metastasis, cellular interactions, signal transduction, phagocytosis, and angiogenesis, among others. At a tisular level, the relationship between proteases and their inhibitors maintains an adjusted balance, which is broken in cases of carcinogenesis, infections and other pathological disorders. Indeed, proteases play a key role in pathogenesis. A large number of physiopathological disorders result from the imbalance of proteolytic activity. In this context protease inhibitors (PIs) are a key component of the endogenous defense system, helping to regulate and balance the cellular proteolytic activities. It has been shown that PIs prevent tumor progression and metastasis and also have the potential to counteract hereditary disorders such as emphysema and epilepsy. PIs may also interfere with the life cycle of many viruses and thus help to prevent some viral diseases. Synthetic PIs are part of a combined therapy against AIDS and have enormous potential for being used against many other diseases. Hence PIs, both natural and synthetic, are a valid way to enrich the pharmacological arsenal existing at the date.Facultad de Ciencias Exacta

Pharmacological potential of plant proteases inhibitors

Las proteasas constituyen un conjunto de enzimas que juegan un papel crucial en numerosos procesos celulares, tales como la degradación de proteínas implicadas en la regulación del ciclo celular, la apoptosis, el crecimiento, la activación, adhesión, invasión y migración celular, las metástasis, las interacciones celulares, la transducción de señales, la fagocitosis y la angiogénesis, entre otros. A nivel tisular la relación entre proteasas y sus inhibidores mantiene un equilibrado balance, equilibrio que se quiebra en casos de carcinogénesis, infecciones y otros desórdenes patológicos. En efecto, las proteasas juegan un papel clave en la patogénesis. Un gran número de desórdenes fisiopatológicos resultan del desbalance de la actividad proteolítica. En este marco los inhibidores de proteasas (IPs) son parte elemental del sistema de defensa endógeno, ya que ayudan a regular y balancear las actividades proteolíticas celulares. Se ha demostrado que los IPs evitan la progresión de tumores y metástasis y también tienen el potencial de contrarrestar desórdenes hereditarios tales como el enfisema y la epilepsia. Los IPs también pueden interferir el ciclo de vida de muchos virus y en consecuencia ayudar a prevenir algunas patologías virales. Por su parte los IPs sintéticos son integrantes de la terapia combinada contra el SIDA y tienen un enorme potencial para ser utilizados contra muchas otras enfermedades. De allí que los IPs, tanto naturales como sintéticos, resultan una alternativa válida para enriquecer el arsenal farmacológico existente a la fecha.Proteases represent an enzyme group that plays a crucial role in many cell processes, such as degradation of proteins involved in the regulation of cell cycle, apoptosis, cell growth and activation; adhesion, invasion, and cell migration; metastasis, cellular interactions, signal transduction, phagocytosis, and angiogenesis, among others. At a tisular level, the relationship between proteases and their inhibitors maintains an adjusted balance, which is broken in cases of carcinogenesis, infections and other pathological disorders. Indeed, proteases play a key role in pathogenesis. A large number of physiopathological disorders result from the imbalance of proteolytic activity. In this context protease inhibitors (PIs) are a key component of the endogenous defense system, helping to regulate and balance the cellular proteolytic activities. It has been shown that PIs prevent tumor progression and metastasis and also have the potential to counteract hereditary disorders such as emphysema and epilepsy. PIs may also interfere with the life cycle of many viruses and thus help to prevent some viral diseases. Synthetic PIs are part of a combined therapy against AIDS and have enormous potential for being used against many other diseases. Hence PIs, both natural and synthetic, are a valid way to enrich the pharmacological arsenal existing at the date.Facultad de Ciencias Exacta

Proteolytic extracts of three bromeliaceae species as eco-compatible tools for leather industry

In addition to the large quantities of solid waste rich in protein, most tanneries use high proportions of Na2S and CaO during the dehairing step, resulting in effluents of high alkalinity and large amounts of suspended solid, besides the risk of liberating the toxic hydrogen sulphide. The current worldwide legislation on environmental requires tanneries to reduce pollution and to replace conventional processes by greener technologies. Enzymes are a technological tool of interest for industry because are able to achieve a high reaction rate under soft pH, temperature, and pressure conditions, besides a high specificity of reaction, biodegradability, non-toxic nature and non-polluting effluent generation. In leather industry enzymes are principally used in pre-tanning operations (soaking, dehairing, bating, and degreasing) and waste treatment. Particularly, proteases have been chosen as a promising eco-friendly alternative to lime and sodium sulphide dehairing. Extracts rich in cysteine proteases with high proteolytic activity (CU) have been obtained from fruits of Bromeliaceae species: Bromelia balansae (Bb), B. hieronymi (Bh), and Pseudananas macrodontes (Pm). In this work, Bb, Bh, and Pm have been studied for application in leather industry compared with commercial enzyme, focusing in their dehairing properties. Enzymatic activities against representative substrates of skin proteins were spectrophotometrically measured at 25, 35, and 55ºC (Tris-HCl, 0.1 M, pH 8, Cys 20 mM). Keratin Azure (KA), Elastin-Congo Red (E), epidermis substrate (EP), and Hide Powder Azure (HPA) were used as representative substrates of keratin, elastin, epidermis, and collagen, respectively. Ability to dehairing was evaluated by incubating soaked cow skins with different concentrations of extracts at 25ºC and pH 8 during 24 h. Grain surface and cross section of skins were observed by scanning electron microscopy. Extracts were able to degrade representative substrates of skin proteins and when compared to the same CU showed similar activity on collagen and epidermis; however, Bh and Pm were the most actives against keratin, while Bh was the only active against elastin. Extracts showing different proteolytic activity (Bb required 1 CU/ml, Bh 1.5 CU/ml, and Pm 0.5 CU/ml) were able to depilate cow skin after a gentle scraping. Although depilated skins with Bb, Bh, and Pm showed different surface aspects, desirable characteristics of dehairing were observed for all extracts since hair pores did not show residual hair, grain surface were clean and intact, and collagen fiber bundles of dermis were not damaged. In conclusion, results here presented show that proteolytic extracts of Bromeliaceae species are promising eco-compatible tools for leather industry, principally in treatment of their waste and dehairing process.Centro de Investigación de Proteínas VegetalesComisión de Investigaciones Científicas de la provincia de Buenos Aire

Proteolytic extracts of three bromeliaceae species as eco-compatible tools for leather industry

In addition to the large quantities of solid waste rich in protein, most tanneries use high proportions of Na2S and CaO during the dehairing step, resulting in effluents of high alkalinity and large amounts of suspended solid, besides the risk of liberating the toxic hydrogen sulphide. The current worldwide legislation on environmental requires tanneries to reduce pollution and to replace conventional processes by greener technologies. Enzymes are a technological tool of interest for industry because are able to achieve a high reaction rate under soft pH, temperature, and pressure conditions, besides a high specificity of reaction, biodegradability, non-toxic nature and non-polluting effluent generation. In leather industry enzymes are principally used in pre-tanning operations (soaking, dehairing, bating, and degreasing) and waste treatment. Particularly, proteases have been chosen as a promising eco-friendly alternative to lime and sodium sulphide dehairing. Extracts rich in cysteine proteases with high proteolytic activity (CU) have been obtained from fruits of Bromeliaceae species: Bromelia balansae (Bb), B. hieronymi (Bh), and Pseudananas macrodontes (Pm). In this work, Bb, Bh, and Pm have been studied for application in leather industry compared with commercial enzyme, focusing in their dehairing properties. Enzymatic activities against representative substrates of skin proteins were spectrophotometrically measured at 25, 35, and 55ºC (Tris-HCl, 0.1 M, pH 8, Cys 20 mM). Keratin Azure (KA), Elastin-Congo Red (E), epidermis substrate (EP), and Hide Powder Azure (HPA) were used as representative substrates of keratin, elastin, epidermis, and collagen, respectively. Ability to dehairing was evaluated by incubating soaked cow skins with different concentrations of extracts at 25ºC and pH 8 during 24 h. Grain surface and cross section of skins were observed by scanning electron microscopy. Extracts were able to degrade representative substrates of skin proteins and when compared to the same CU showed similar activity on collagen and epidermis; however, Bh and Pm were the most actives against keratin, while Bh was the only active against elastin. Extracts showing different proteolytic activity (Bb required 1 CU/ml, Bh 1.5 CU/ml, and Pm 0.5 CU/ml) were able to depilate cow skin after a gentle scraping. Although depilated skins with Bb, Bh, and Pm showed different surface aspects, desirable characteristics of dehairing were observed for all extracts since hair pores did not show residual hair, grain surface were clean and intact, and collagen fiber bundles of dermis were not damaged. In conclusion, results here presented show that proteolytic extracts of Bromeliaceae species are promising eco-compatible tools for leather industry, principally in treatment of their waste and dehairing process.Centro de Investigación de Proteínas VegetalesComisión de Investigaciones Científicas de la provincia de Buenos Aire

Proteolytic extracts of three bromeliaceae species as eco-compatible tools for leather industry

In addition to the large quantities of solid waste rich in protein, most tanneries use high proportions of Na2S and CaO during the dehairing step, resulting in effluents of high alkalinity and large amounts of suspended solid, besides the risk of liberating the toxic hydrogen sulphide. The current worldwide legislation on environmental requires tanneries to reduce pollution and to replace conventional processes by greener technologies. Enzymes are a technological tool of interest for industry because are able to achieve a high reaction rate under soft pH, temperature, and pressure conditions, besides a high specificity of reaction, biodegradability, non-toxic nature and non-polluting effluent generation. In leather industry enzymes are principally used in pre-tanning operations (soaking, dehairing, bating, and degreasing) and waste treatment. Particularly, proteases have been chosen as a promising eco-friendly alternative to lime and sodium sulphide dehairing. Extracts rich in cysteine proteases with high proteolytic activity (CU) have been obtained from fruits of Bromeliaceae species: Bromelia balansae (Bb), B. hieronymi (Bh), and Pseudananas macrodontes (Pm). In this work, Bb, Bh, and Pm have been studied for application in leather industry compared with commercial enzyme, focusing in their dehairing properties. Enzymatic activities against representative substrates of skin proteins were spectrophotometrically measured at 25, 35, and 55ºC (Tris-HCl, 0.1 M, pH 8, Cys 20 mM). Keratin Azure (KA), Elastin-Congo Red (E), epidermis substrate (EP), and Hide Powder Azure (HPA) were used as representative substrates of keratin, elastin, epidermis, and collagen, respectively. Ability to dehairing was evaluated by incubating soaked cow skins with different concentrations of extracts at 25ºC and pH 8 during 24 h. Grain surface and cross section of skins were observed by scanning electron microscopy. Extracts were able to degrade representative substrates of skin proteins and when compared to the same CU showed similar activity on collagen and epidermis; however, Bh and Pm were the most actives against keratin, while Bh was the only active against elastin. Extracts showing different proteolytic activity (Bb required 1 CU/ml, Bh 1.5 CU/ml, and Pm 0.5 CU/ml) were able to depilate cow skin after a gentle scraping. Although depilated skins with Bb, Bh, and Pm showed different surface aspects, desirable characteristics of dehairing were observed for all extracts since hair pores did not show residual hair, grain surface were clean and intact, and collagen fiber bundles of dermis were not damaged. In conclusion, results here presented show that proteolytic extracts of Bromeliaceae species are promising eco-compatible tools for leather industry, principally in treatment of their waste and dehairing process.Centro de Investigación de Proteínas VegetalesComisión de Investigaciones Científicas de la provincia de Buenos Aire

Estudio preliminar de "Macrodontina", una nueva cisteinilproteinasa aislada de frutos de Pseudananas macrodontes (Morr.) Harms (Bromeliaceae)

From inmature fruits of Pseudananas macrodontes (Morr.) Harms, known as ihvirá in the northeast of Argentina, has been isolated and partially characterized a sulfhydril-protease highly active between pH 7 and 10. The acetonic precipitates are quite stable at temperatures below 45ºC and not affected by freezing or liophylization. Anionic exchange chromatography (DEAE-Sepharose Fast Flow) affords two active fractions, both well resolved by gel filtration (Sephadex G-75). The electrophoretic pattern (SDS-PAGE) of the acetonic precipitate shows six protein fractions, between 14 and 67 kDa, but the zymogram reveals the presente of three active fractions, of about 27,30, arid 54 kDa.De los frutos semimaduros del ihvirá (Pseudananas macrodontes (Morr.) Harms (Bromeliaceae) se ha logrado aislar y caracterizar parcialmente una proteasa sulfhidrílica ( macrodontina ) con actividad destacada entre pH 7 y 10. Las preparaciones parcialmente purificadas (precipitados acetónicos) son relativamente estables a temperaturas inferiores a 45ºC y no se alteran por congelamiento o liofilización. Una ulterior purificación por cromatografía de intercambio aniónico (DEAE-Sepharose Fast Flow) permite detectar dos fracciones activas, que son bien resueltas por cromatografía de exclusión molecular (Sephadex G-75). Por electroforesis (SDS-PAGE) del precipitado acetónico se detectan seis fracciones proteicas (entre 14 y 67 kDa), pero el zimograma revela que sólo tres de ellas tienen actividad caseinolítica, de pesos moleculares de alrededor de 27,30 y 54 kDa

Evaluation of Anti-Inflammatory Activity of Pseudananas macrodontes (Morr.) Harms (Bromeliaceae) Fruit Extract in Rats

Several species of the family Bromeliaceae are characterized by the production of proteases in unusual amounts, especially in fruits. Bromelain, an extract rich in cysteine endo-peptidases obtained from Ananas comosus L., and a few other proteases have been used as anti-infl ammatory agents for some years, but bromelain is still mainly being used as alternative and/or complementary therapy to the treatment with glucocorticoids, nonsteroi-dal antirheumatics, and immunomodulators. In this study, the anti-infl ammatory action of a partially purifi ed extract from Pseudananas macrodontes (Morr.) Harms fruits (PPEPm) is presented, whose main components are cysteine endopeptidases. The effect of PPEPm was assessed in carrageenan-induced and serotonin-induced rat paw edema, as well as in the cotton pellet granuloma model. Doses with equal proteolytic activity of PPEPm and bromelain produced signifi cantly similar anti-infl ammatory responses in the acute infl ammatory models assayed, supporting the hypothesis that proteolytic activity could be responsible for the anti-infl ammatory action. On the contrary, comparable anti-infl ammatory effects of PPEPm and bromelain in the chronic infl ammatory assay required a much lower proteolytic activity content of PPEPm, which could be due to a differential affi nity for the protein target involved in this process.Facultad de Ciencias Exacta

Caracterización parcial de una proteinasa coagulante de la leche aislada del alcaucil (Cynara scolymus L., Asteraceae)

The presence of proteinases in Cynara scolyrnus L. ( artichoke ) has been investigated by determining the proteolytic and milk clotting activities of crude extracts of different parts of the inflorescence in various stages of development, as well as of leaves and roots. Although all the preparations showed a certain extent of proteolytic activity, only those of adult leaves, pappus, and immature and mature flowers were able to clot milk. The extract of the upper (violet) part of mature flowers exhibited optimum activity at acid pH values (90% of maximum activity at pH 3.5 - 5.0) which was strongly inhibited by pepstatine A, suggesting the presence of aspartic proteinases. This extract had a low thermal stability at temperatures above 45ºC, which could be a useful property in cheese making process, as it could be quickly inactivated by moderate heating.Se ha estudiado la presencia de proteinasas en el alcaucil (Cynara scolymus L.) midiendo la actividad proteolítica y la capacidad coagulante de la leche de preparaciones crudas de diferentes partes de la inflorescencia en distintos estadios de desarrollo, así como en raíces y hojas. Si bien se detecta actividad proteolitica en todas las preparaciones, sólo las de hojas adultas, de papus y de flores inmaduras y maduras son capaces de coagular la leche. El extracto de la parte superior (violeta) de las flores maduras exhibe un perfil de pH que es óptimo en la zona ácida (90% de máxima actividad entre pH 3,5 y 5,0), pero la actividad es fuertemente inhibida por pepstatina A, lo que sugeriría la presencia de una o más proteinasas aspárticas. La estabilidad térmica del extracto es baja a temperaturas superiores a 45ºC, circunstancia que puede resultar de utilidad en la producción de quesos, ya que la enzima puede ser inactivada a temperaturas moderadas

Proteolytic properties of <i>Funastrum clausum</i> latex

As part of a screening of latex endopeptidases from plants growing in Argentina, the presence of proteolytic activity in the latex of Funastrum clausum stems is reported. The proteases present in the crude extract showed the main characteristics of the cysteine proteolytic class, i.e. optimum pH at alkaline range, isoelectric point (pI) higher than 9.0, and inhibition of proteolytic activity by thiol blocking reagents. A remarkable thermal stability was also evident in the crude extract. Endosterolytic preference tried on p-nitrophenyl esters of N-a-carbobenzoxy-L-amino acids was higher for the alanine, asparagine and tyrosine derivatives. Preliminary peptidase purification by two-step ionic exchange showed the presence of two proteolytic fractions with molecular masses of approximately 24.0 kDa according to SDS-PAGE.Centro de Investigación de Proteínas Vegetale