Assay of Proteolytic Enzymes

Davis, Neil C.; Smith, Emil L.

doi:10.1002/9780470110188.ch8

Cited by 61 publications

(8 citation statements)

References 57 publications

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“…Free proline content was extracted from leaves in 3 % aqueous sulphosalicylic acid and estimated using ninhydrin reagent (Bates et al 1973). The content of chlorophyll (Chl) and protease activity were determined by the methods of Lichtenthaler (1987) and Davis and Smith (1955), respectively. The assay for ribulose-1,5-bisphosphate carboxylase (RuBPC) and NR activities were carried out as described by Nedunchezhian and Kulandaivelu (1991).…”

mentioning

confidence: 99%

Effect of water deficit on photosynthetic and other physiological responses in grapevine (Vitis vinifera L. cv. Riesling) plants

Bertamini¹,

Zulini²,

Muthuchelian³

et al. 2006

Photosynt.

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The grapevine (Vitis vinifera L. cv. Riesling) plants subjected to water deficit were studied for changes in relative water content (RWC), leaf dry mass, contents of chlorophyll (Chl), total leaf proteins, free amino acids, and proline, and activities of ribulose-1,5-bisphosphate carboxylase (RuBPC), nitrate reductase (NR), and protease. In water-stressed plants RWC, leaf dry matter, Chl content, net photosynthetic rate (P N ), and RuBPC and NR activities were significantly decreased. The total leaf protein content also declined with increase in the accumulation of free amino acids. Concurrently, the protease activity in the tissues was also increased. A significant two-fold increase in proline content was recorded.Additional key words: amino acids; chlorophyll; net photosynthetic rate; nitrate reductase; proline; protease; relative water content; ribulose-1,5-bisphosphate carboxylase.---Water stress has a multifaceted effect on plant growth and metabolism (Hsiao 1973). Under stress, leaves close their stomata, and this is generally believed to be the cause of reduced carbon gain under periods of drought (Kaiser et al. 1981). Water stress affects not only the carbon metabolism but also nitrogen metabolism. During water stress nitrate reductase (NR) activity is lost more rapidly than most enzymes (Huffaker et al. 1970). The other enzymes of the pathway of nitrate assimilation, nitrite reductase and glutamine synthetase (Taylor et al. 1982), are relatively unaffected. Huffaker et al. (1970) suggested that a rapid loss of NR activity could be part of a biochemical adaptation to water deficit; shutting off the nitrate assimilation pathway at the first step would reduce energy requirements during periods of stress and prevent accumulation of nitrite and ammonium.Water stress is the most important factor limiting grapevine growth in the Mediterranean area (Gomez del Campo et al. 2000, Flexas et al. 2002. Previous studies on the photosynthetic response to drought under field conditions showed that stomatal closure is an early response, which is almost matched by decreases in CO 2 assimilation. This response becomes progressively greater through summer, as soil water availability decreases and also non-stomatal effects appear (Escalona et al. 1997). The purpose of the present study was the investigation of the effects of water deficit on physiological responses in grapevine.One-year-old grapevine (Vitis vinifera L. cv. Riesling) plants were grown in 20 000-cm 3 pots containing soil : sand : peat : vermiculite (3 : 1 : 3 : 3) in glasshouse [26/30-16/20 o C day/night temperature, photosynthetic photon flux density (PPFD) of 1 200-1 500 µmol m -2 s -1 , 14/10 h light/dark cycle, and relative humidity between 65 and 70 %], and then divided into two uniform groups of ten pots each. The first group (control) continued to receive daily irrigation in order to maintain the soil water at the field capacity while in the second group (water deficit) irrigation was stopped. Measurements were made 10 d after the irrigation was stopped...

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mentioning

confidence: 99%

Effect of water deficit on photosynthetic and other physiological responses in grapevine (Vitis vinifera L. cv. Riesling) plants

Bertamini¹,

Zulini²,

Muthuchelian³

et al. 2006

Photosynt.

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“…The free amino acid content was estimated by the method of Moore and Stein (1954) as described by Colowick and Kaplan (1957) and the protease activity by the method of Davis and Smith (1955). The activity levels of aspartate aminotransferase (AAT) and alanine aminotransferase (AlAT) were estimated by the method of Reitman and Frankel (1957) as described by Bergmeyer and Bruns (1965) and the activity of glutamate dehydrogenase (GDH) was assayed by the method of Lee and Lardy (1965).…”

Section: Methodsmentioning

confidence: 99%

Effect of ultrasound on protein metabolism in the silkworm, <i>Bombyx mori</i> (L.)

Murali¹,

P²,

Sahitya³

2015

Eth J Sci & Technol

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The parameters of protein metabolism, such as the levels of soluble, structural and total proteins, free amino acids, and the activity levels of protease, aspartate and alanine aminotransferases and glutamate dehydrogenase were assayed in the hemolymph, silk-gland, muscle and fat-body on different days of the 5 th instar larva of the silkworm, Bombyx mori, following exposure of the silkworm eggs to 1 MHz continuous wave of ultrasound at an intensity of 9W/cm 2 for 2 minutes. Ultrasound was found to promote the accumulation of proteins, which include silk proteins as well, while retarding proteolysis and turnover of proteins towards the release of amino acids, keto-acids etc. Changes in the levels of these biochemical constituents are correlated with the events of histogenesis and histolysis associated with metamorphosis. It may be inferred that protein metabolism is stimulated by ultrasound, resulting in greater turnover of silk proteins, spinning activity and silk output.

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“…The contents were centrifuged at 2,500 rpm for 10 minutes in a refrigerated centrifuge, and the cell-free extracts were used for enzyme assays. Protease activity was measured (Davis and Smith, 1955) with the reaction mixture containing 100 mL of phosphate buffer (pH 5.0) and 12 mg of denatured protein. AST and ALT activities were assayed following the method of Reitman and Frankel (1957).…”

Section: Biochemical Estimationsmentioning

confidence: 99%

Modulation in the protein metabolism by subacute sodium cyanide intoxication in the freshwater fish,Labeo rohita(Hamilton)

Dube¹,

Hosetti²

2011

Drug and Chemical Toxicology

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The effects of exposure to one-third and one-fifth sublethal concentrations (0.106 and 0.064 mg/L) of sodium cyanide on protein metabolism on freshwater carp, Labeo rohita, was studied. Three functionally different tissues, namely, the liver, muscle, and gills, were studied after 5, 10, and 15 days. Exposures produced marked changes in protein metabolic profile in all tissues studied. These changes were more pronounced in the one-third sublethal concentration, suggesting a cumulative action of toxicant. This investigation revealed that the total, structural, and soluble proteins and urea content in all the three tissues were decreased, whereas free amino acids, ammonia, and enzyme activity (i.e., protease, alanine aminotransferase, and aspartate aminotransferase) exhibited elevated levels at both sublethal concentrations. Variation in protein metabolism in the fish, induced by sodium cyanide, demonstrated its toxic effects on cellular metabolism, thereby leading to impaired protein synthetic machinery. The results of the present study indicate that a mechanism of impaired energy transformation has direct action on the fish, L. rohita, and its impact is clearly evident from the change in the nutritional content of the fish.

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Assay of Proteolytic Enzymes

Cited by 61 publications

References 57 publications

Effect of water deficit on photosynthetic and other physiological responses in grapevine (Vitis vinifera L. cv. Riesling) plants

Effect of water deficit on photosynthetic and other physiological responses in grapevine (Vitis vinifera L. cv. Riesling) plants

Effect of ultrasound on protein metabolism in the silkworm, <i>Bombyx mori</i> (L.)

Modulation in the protein metabolism by subacute sodium cyanide intoxication in the freshwater fish,Labeo rohita(Hamilton)

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