I intend to purify the enzymes from the crude enzyme extract. The extract contains at least five enzymes with various molecular weights and net charges at pH 7: Molecular Weight (Da) Net charges at pH 7 Proteins/enzymes 48,500 5,000 250,000 150,000 A В ++ C D 60,500 aArrange the elution profile sequence of these enzymes in a Gel Filtration 'Chromatography from the first eluted to the last. Justify your answer. Arrange the elution profile sequence of these proteins in an Anion Exchange Chromatography from the first eluted to the last. Justify your E +
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Idris has successfully extracted enzymatic proteins from the fish viscera (intestines and stomach). After homogenization and centrifugation, he managed to pool the crude enzyme extract. He is characterizing the enzymes. Please help Idris by answering the following
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- To purify a given enzyme from a crude extract that contains it, fractional precipitations, exchange chromatography are successively tested ionic and exclusion chromatography, with the results shown in the following table: Volume of dissolution Protein concentration Enzymatic activity (ml) (mg/ml) (U/ml) a) Calculate the percentage of recovery of the enzyme after each of the manipulations to which it has been subjected. b) Indicate if it is necessary to continue purifying the enzyme or if it is expected to have reached its electrophoretic homogeneity.A biochemist discovers and purifies a new enzyme, generating the purification table below. Procedure Total Protein (Mg) Activity (Units) Crude Extract 20,000 4,000,000 Precipitation (Salt) 5,000 3,000,000 Precipitation (pH) 4,000 1,000,000 Ion Exchange Chromatography 200 800,000 Affinity Chromatography 50 750,000 Size-exclusion Chromatography 45 675,000 a) From the information given in the table, calculate the specific activity of the enzymeafter each purification procedure.b) Which of the purification procedures used for this enzyme is most effective (i.e., givesthe greatest relative…200 ml of a 2% protein solution containing an enzyme that you want to purify. Half of the sample is subjected to method A, consisting of fractionated precipitations and 5 ml of final solution are obtained, with a concentration protein equal to 5 mg / ml and enzymatic activity equal to 2000 U / ml. The other half is subjected to method B, consisting of ion exchange chromatography, and a final solution of 10 ml, with protein richness equal to 10 mg / ml and with an activity enzymatic also equal to 2000 U / ml. You want to know: a) Which of the two methods has provided the purest enzyme. b) By which of the methods the greatest amount of enzyme has been obtained.
- An enzyme is present at 100 nM (nanomolar) and has a Vmax value of 25 uM/s (micromolar/second). The Km for the substrate is 5.2 uM. What is the kcat value? Report your answer to three significant figures in units of 1/s.An enzyme catalysed reaction has a Km of 8 mM and a Vmax of 13 nM.s-1. Use the Michaelis-Menten equation to calculate the reaction velocity when the substrate concentration is 18 mM.You need a buff er at pH 7.5 for use in purifying a protein at 4°C. You have chosen Tris, pK 8.08, ΔH° = 50 kJ · mol−1. You carefully make up 0.01 M Tris buffer, pH 7.5 at 25°C, and store it in the cold to equilibrate it to the temperature of the purifi cation. When you measure the pH of the temperature-equilibrated buff er it has increased to 8.1. What is the explanation for this increase? How can you avoid this problem?
- True or False Immobilization improves the stability of the enzyme. EnaLne, has a half-life of 10 days in free solution, but under identical conditions of temperature, pH, and medium composition, the measured half-life of a packed column is 30 days. The enzyme is immobilized in a porous sphere 5 mm in diameter.Aerobic degradation of an organic compound by mixed cultureof organism in wastewater can be represented by following reaction. C3H6O3 + a O2 + b NH3 → c C5H7NO2 + d H2o + e CO2 A. Determine a, b, c, d and e, if YX/S = 0.4 d X/g S. B. Determine the yield coefficients YX/O2 and YX/NH3. C. Determine the degree of reductions for the substrate, bacteria and RQ for the organismsQuantitative Estimation of Amino Acids by Ninhydrin http://vlab.amrita.edu/?sub=3&brch=63&sim=156&cnt=2 can u help me with question 2 of the assignment questions Based on the experimental data provided, estimate the amount of amino acid in the given unknown solution by Ninhydrin method. SI No. Volume of standard amino acid solution (ml) Amount of amino acid (µg) OD at 570nm 1 Blank 0 2 0.2 0.12 3 0.4 0.25 4 0.6 0.45 5 0.8 0.55 6 1.0 100 0.68 7 Unknown (0.5ml) 0.41
- You are given as following : 20 µl pure LDH on ice, 2.0ml of 6mM NAD+, 2.0ml of 150mM lactate, and 0.14M CAPS buffer. LDH reaction cocktail has final concentration of 1mM NAD+ and 25mM lactate in 0.14M CAPS buffer. LDH activityis measured by mixing 10 µl of LDH sample and 990 µl LDH reaction cocktail before getting ∆A340/min reading on spectrometer. (a). Describe in detail how you would prepare for your LDH reaction cocktail including how to make dilutions.33333333333333333333333 Using the table below, differentiate the effect of two varying pH levels (as indicated by by the color) to the amylase enzyme. How does pH level affects the enzymatic reaction (enzyme-substrate complex)? Tube 1 Tube 2 Tube 3 Tube 4 Ingredients StarchAmylaseBuffer pH 7 StarchAmylaseBuffer pH 2 MaltoseWaterBuffer pH 7 StarchWaterBuffer pH 7 Color (1) (2) Orange BlueCalculate your dilution strategyto make a 625 ng/mLHRP solution from a 12.5 μg/mL stock given the totalvolume of enzyme youwillneed for your condition this week. Note if you are testing pH, you only need to calculate one enzyme dilution because all other pHs will use the same strategy.