Activity of Subtilisin Carlsberg in macromolecular crowding

Shaw, Ajay Kumar; Pal, Samir Kumar

doi:10.1016/j.jphotobiol.2006.10.001

Cited by 24 publications

(40 citation statements)

References 33 publications

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“…The signal has been found to be highly monodispersed, and the measured hydrodynamic diameter of 4.3 ( 0.4 nm is consistent with previous studies. 29 It is evidenced from the figure that with an increase in temperature, the micellar diameter changes marginally. The high monodispersity of the DLS signal is retained in all the studied temperatures (figures not shown).…”

Section: Methodsmentioning

confidence: 92%

Temperature-Dependent Hydration at Micellar Surface: Activation Energy Barrier Crossing Model Revisited

2007

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In recent years, the validity of the activation energy barrier crossing model at the micellar surface brings notable controversy (Sen, P.; Mukherjee, S.; Halder, A.; Bhattacharyya, K. Chem. Phys. Lett. 2004, 385, 357-361. Kumbhakar, M.; Pal, H. J. Phys. Chem. B 2004, 108, 19246-19254.) in the literature. In order to check the validity of the model by time-resolved solvation of a probe fluorophore, a wider range of temperature must be considered. At the same time, spatial heterogeneity (solubilization) of the probe and structural perturbation of the host micelle should carefully be avoided, which was not strictly maintained in the earlier studies. We report here the solvation dynamics of 4-(dicyanomethylene)-2-methyl-6(p-dimethylamino-styryl) 4H-pyran (DCM) in the SDS micelle at 298, 323, and 348 K. The probe DCM is completely insoluble in bulk water in this wide range of temperature. The size of the micelle at different temperatures using the dynamic light scattering (DLS) technique is found to have insignificant change. The hydration number of the micelle, determined by sound velocity measurements, decreases with increasing temperature. Time-resolved fluorescence anisotropy reveals the retention of the probe in the micellar interface within the temperature range. The average solvation time decreases with increasing temperature. The result of the solvation study has been analyzed in the light of energetics of bound to free water conversion at a constant size and decreasing hydration number at the micellar surface. The solvation process at the micellar surface has been found to be the activation energy barrier crossing type, in which interfacially bound type water molecules get converted into free type molecules. We have calculated E a to be 3.5 kcal mol -1 , which is in good agreement with that obtained by molecular dynamics simulation studies.

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Section: Methodsmentioning

confidence: 92%

Temperature-Dependent Hydration at Micellar Surface: Activation Energy Barrier Crossing Model Revisited

2007

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“…For instance, the activity and specificity of KLK3 [34], which is expressed only in the prostate, is highly influenced by the sodium citrate concentration, and prostate contains the highest concentration of this anion in the body [68]. The macromolecular crowding concept seems to be adequate to interpret the high sodium citrate and sulfate concentrations that compact the proteins with reduction of surface-to-volume ratio, which results in an active conformation of the enzyme; for review and concept explanation of macromolecular crowding see [69][70][71] and for proteases other than kallikreins see [72,73].…”

Section: Discussionmentioning

confidence: 99%

Specificity studies on Kallikrein-related peptidase 7 (KLK7) and effects of osmolytes and glycosaminoglycans on its peptidase activity

Oliveira

Bertolin

Andrade

et al. 2015

Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics

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“…The serine protease activity was assessed via the previously described method [12], with slight modifications, using the peptidyl fluorogenic substrate, AAF-AMC. In brief, 10μl of purified enzyme was preincubated for 2 h at 37°C with 85μl of 100 mM Tris buffer/pH7.5 and 2 mM dithiothreitol (DTT).…”

Section: Enzyme Activity Assaymentioning

confidence: 99%

Purification, Molecular Cloning, and Biochemical Characterization of Subtilisin JB1 from a Newly Isolated Bacillus subtilis JB1

Sung

Ahn

Kim

et al. 2009

Appl Biochem Biotechnol

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An extracellular gelatinolytic enzyme obtained from the newly isolated Bacillus subtilis JB1, a thermophilic microorganism relevant to the aerobic biodegradation process of fish-meal production, was purified via ammonium sulfate precipitation, Sephadex G-200 Gel filtration chromatography, and one-dimensional gel electrophoresis separation and subsequently identified via peptide mass fingerprinting and chemically assisted fragmentation matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The subtilisin JB1 gene was sequenced and its recombinant protein prosubtilisin JB1 was expressed in Escherichia coli, and the purified prosubtilisin JB1 (62 kDa) protein was digested with gelatin, bovine serum albumin, azocasein, fibrinogen, and the fluorogenic peptide substrate Ala-Ala-Phe-7-amido-4-methylcoumarin hydrochloride, whereas the serine protease inhibitors phenylmethylsulfonyl fluoride and chymostatin completely inhibited its enzyme activity at an optimal pH of 7.5. Thus, our results show that subtilisin JB1 may serve as a potential source material for use in industrial applications of proteolytic enzymes and microorganisms for fishery waste degradation and fish by-product processing.

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Activity of Subtilisin Carlsberg in macromolecular crowding

Cited by 24 publications

References 33 publications

Temperature-Dependent Hydration at Micellar Surface: Activation Energy Barrier Crossing Model Revisited

Temperature-Dependent Hydration at Micellar Surface: Activation Energy Barrier Crossing Model Revisited

Specificity studies on Kallikrein-related peptidase 7 (KLK7) and effects of osmolytes and glycosaminoglycans on its peptidase activity

Purification, Molecular Cloning, and Biochemical Characterization of Subtilisin JB1 from a Newly Isolated Bacillus subtilis JB1

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