Chapter 8 Quantitation of Protein

Noble, James; Bailey, Marc J. A.

doi:10.1016/s0076-6879(09)63008-1

Cited by 418 publications

(270 citation statements)

References 35 publications

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“…The purified protein was concentrated to ϳ10 mg/ml using a Millipore Ultrafree device with a molecular mass cutoff of 10,000 Da. Typical yields of purified protein, as determined by Bradford assay (14), were 2-4.5 mg/liter of culture. Protein was characterized by activesite titration using Z-VAD-FMK and found to be 100% active.…”

Section: Methodsmentioning

confidence: 99%

Structural and Enzymatic Insights into Caspase-2 Protein Substrate Recognition and Catalysis

Tang

Wells

Arkin

2011

Journal of Biological Chemistry

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Caspase-2, the most evolutionarily conserved member in the human caspase family, may play important roles in stress-induced apoptosis, cell cycle regulation, and tumor suppression. In biochemical assays, caspase-2 uniquely prefers a pentapeptide (such as VDVAD) rather than a tetrapeptide, as required for efficient cleavage by other caspases. We investigated the molecular basis for pentapeptide specificity using peptide analog inhibitors and substrates that vary at the P5 position. We determined the crystal structures of apo caspase-2, caspase-2 in complex with peptide inhibitors VDVAD-CHO, ADVAD-CHO, and DVAD-CHO, and a T380A mutant of caspase-2 in complex with VDVAD-CHO. Two residues, Thr-380 and Tyr-420, are identified to be critical for the P5 residue recognition; mutation of the two residues reduces the catalytic efficiency by about 4-and 40-fold, respectively. The structures also provide a series of snapshots of caspase-2 in different catalytic states, shedding light on the mechanism of capase-2 activation, substrate binding, and catalysis. By comparing the apo and inhibited caspase-2 structures, we propose that the disruption of a non-conserved salt bridge between Glu-217 and the invariant Arg-378 is important for the activation of caspase-2. These findings broaden our understanding of caspase-2 substrate specificity and catalysis.

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Section: Methodsmentioning

confidence: 99%

Structural and Enzymatic Insights into Caspase-2 Protein Substrate Recognition and Catalysis

Tang

Wells

Arkin

2011

Journal of Biological Chemistry

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“…Homogenates were then centrifuged at 12000 × g for 10 min at 4 °C. The supernatants was collected and stored until use at -80 o C. Protein concentration was determined by Bradford's method (Noble, Bailey, 2009). …”

Section: Animalsmentioning

confidence: 99%

Hepatoprotective effect of Phytosome Curcumin against paracetamol-induced liver toxicity in mice

Tùng¹,

Hải

Phan

2017

Braz. J. Pharm. Sci.

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Curcuma longa, which contains curcumin as a major constituent, has been shown many pharmacological effects, but it is limited using in clinical due to low bioavailability. In this study, we developed a phytosome curcumin formulation and evaluated the hepatoprotective effect of phytosome curcumin on paracetamol induced liver damage in mice. Phytosome curcumin (equivalent to curcumin 100 and 200 mg/kg body weight) and curcumin (200 mg/kg body weight) were given by gastrically and toxicity was induced by paracetamol (500 mg/kg) during 7 days. On the final day animals were sacrificed and liver function markers (ALT, AST), hepatic antioxidants (SOD, CAT and GPx) and lipid peroxidation in liver homogenate were estimated. Our data showed that phytosome has stronger hepatoprotective effect compared to curcumin-free. Administration of phytosome curcumin effectively suppressed paracetamol-induced liver injury evidenced by a reduction of lipid peroxidation level, and elevated enzymatic antioxidant activities of superoxide dismutase, catalase, glutathione peroxidase in mice liver tissue. Our study suggests that phytosome curcumin has strong antioxidant activity and potential hepatoprotective effects.Uniterms: Curcumin/effects. Curcumin/antioxidant activity. Curcuma longa. Phytosome. Hepatoprotective.

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“…A given volume of Tris-buffered saline was added to a tissue sample while stirring in ice bath. The content was centrifuged at 4 °C for ten minutes at 1000 x g. The supernatant was then separated and its total protein content, expressed in milligrams of protein, was determined by using Bradford's method (Noble and Bailey, 2009). …”

Section: Tissue Sample Preparationmentioning

confidence: 99%

Effect of Curcumin on Oxidative Stress in a Model of Turpentine Induced Acute Experimental Inflammation

Coneac

Orăsan

Leucuţa

et al. 2017

Not Bot Horti Agrobo

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Curcumin, a natural phenolic compound is an anti-tumor agent with anti-inflammatory and anti-oxidant properties. The aim of this research was to evaluate oxidative stress levels, the antioxidant activity and Curcumin concentrations by high performance liquid chromatography (HPLC) in an acute experimental inflammation induced by Turpentine oil (intramuscular 0.6 mg kg -1 body weight) and to compare a prophylactic versus a therapeutic regimen of Curcumin (oral suspension of 150 mg Curcumin kg -1 rat weight). Sixteen adult male Wistar rats were assigned to four groups: Control, Group I (Curcumin only), Group II (Curcumin administration, then induced inflammation after 1 hour) and Group III (induced inflammation then Curcumin administration after 2 hours). Oxidative stress was assessed by measuring serum malondialdehide and carbonylated proteins, while systemic and local total antioxidant capacity was determined by ABTS. Local tissue changes (muscle, kidney, liver) were analysed using histopathology. Results showed that acute inflammation significantly increased lipid peroxidation in Groups II and III compared to Control and Group I. A significantly reduced total antioxidant capacity (ATBS) was present in serum and kidney in Group II, also in muscle and kidney in Group III. ABTS levels were significantly increased only in the liver tissue of the animals in Groups II and III with induced inflammation as compared to Group I. This study proved the potential of Curcumin in reducing oxidative stress in both prophylactic and therapeutic regimens.

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Chapter 8 Quantitation of Protein

Cited by 418 publications

References 35 publications

Structural and Enzymatic Insights into Caspase-2 Protein Substrate Recognition and Catalysis

Structural and Enzymatic Insights into Caspase-2 Protein Substrate Recognition and Catalysis

Hepatoprotective effect of Phytosome Curcumin against paracetamol-induced liver toxicity in mice

Effect of Curcumin on Oxidative Stress in a Model of Turpentine Induced Acute Experimental Inflammation

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