### Enzyme Kinetics of Sphingosine KinaseSphingosine-1-phosphate (SPP) has recently been discovered to be important for cell survival. The synthesis of SPP from sphingosine and ATP is catalyzed by the enzyme sphingosine kinase. An understanding of the kinetics of the sphingosine kinase reaction may be important in the development of drugs to treat cancer. The velocity of the sphingosine kinase reaction was measured in the presence (5 μM) and absence of **threo-sphingosine**, a stereoisomer of sphingosine. The results are shown below. The sphingosine kinase used in all the reactions was 10 mg.#### Kinetic Data Table:| Sphingosine (μM) | v₀ (mg·min⁻¹) (no inhibitor) | v₀ (mg·min⁻¹) (with **threo-sphingosine**) ||------------------|-----------------------------|-----------------------------------------|| 2.5 | 32.3 | 7.6 || 3.5 | 40 | 10.5 || 5 | 50.8 | 14.6 || 10 | 72 | 25.4 || 20 | 87.7 | 43.9 || 50 | 115.4 | 70.8 |#### Analytical Questions:1. **What are the KM and VMAX in the presence and absence of the inhibitor?** - To determine the KM and VMAX, Michaelis-Menten plots or Lineweaver-Burk plots of the data would be used.2. **What kind of an inhibitor is threo-sphingosine? What is the KI value?** - Identify whether threo-sphingosine is a competitive, noncompetitive, or uncompetitive inhibitor by analyzing the impact on KM and VMAX. - KI value can be calculated using inhibitor kinetics equations.3. **What is the Kcat (turn over number)?** - The K<

For a one-substrate enzyme-catalyzed reaction, the Michaelis-Menton equation shows the quantitative…

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Biochemistry

9th Edition

ISBN:9781319114671

Author:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.

Publisher:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.

Chapter1: Biochemistry: An Evolving Science

Section: Chapter Questions

Problem 1P

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The active site of an enzyme that uses a general acid-base catalytic mechanism contains a Glu and an Asp residue (both of which are essential for catalysis) with pKa values of 5.9 and 4.5, respectively. If the enzyme is found in the lysosome (pH = 5.2), which residue will act as the general acid and which will act as the general base during the initial steps of the reaction?
The first order decomposition of cyclobutane, C4H8 is shown below: C4H8(g) 2 C2H4(g) The value of the rate constant is 9.20 x 10-3 s-1 at 500°C. Initially, 100.0 kg of C4H8 is present in a container. How long will it take for 80.0 kg of C4Hg to decompose?
With the help of the half reactions given in Table 1, formulate the redox equation for the oxidation of succinate and reduction of ubiquinone.
A particular enzyme-catalyzed reaction has an apparent Vmax = 9.00 nmol s-1 and α' = 3.00 when 2.00 µmol L-1 inhibitor X is present and uncompetitively inhibiting the reaction. Calculate Vmax for the uninhibited reaction in nmol s-1.
One of your colleagues has obtained a sample of muscle phosphorylase b that is known to be relatively inactive. She has approached you for advice on how to set up an appropriate assay. She has the following items available, not all of which are appropriate for this study. Help her out by selecting the items that she should use and what their purpose is in the assay, and then explain why each of the other items would not be useful. 1. 100 UM AMP 2. 100 UM GTP 3.100 uM glucose 4. 100 uM glucose 6-phosphate 5. Branched glycogen 6. Amylose (i.e. unbranched glycogen) 7.50 mM HEPES buffer, pH 7.5 8. 50 mM potassium phosphate buffer, pH 7.5 Write out a short explanation for each of the above items, and upload your answers by the due date.
Allosteric inhibitors of hemoglobin will decrease enzyme activity (oxygen binding) through which of the following mechanisms? Binding the enzyme in "T" conformation and displacing an activator Binding the enzyme and enhancing the Vmax Binding the enzyme, and keeping it in the "R" conformation Binding the enzyme, and keeping it in the "T" conformation
In a major metabolic pathway involving the monosaccharide glucose, one of the reactions involve the conversion of glucose to glucose-6-phosphate summarized below together with accompanying free energy change: glucose + phosphate = glucose-6-phosphate + H,0 AG = 13.8 kJ · mol-1 (Reaction 1) In cells, the production of G6P (Reaction 1 above) is coupled to a reaction that involves the hydrolysis of ATP to ADP (shown below, Reaction 2) which makes the overall reaction much more favorable to the production of glucose-6-phosphate. ATP + H,O = ADP + phosphate (Reaction 2) Why do you think coupling the production of glucose-6-phosphate to the hydrolysis of ATP makes the overall reaction spontaneous? What can you say about the free energy change accompanying the hydrolysis of ATP (Reaction 2)?
During glycogen synthesis, glucose-1P is converted into a molecule called UDPG. This reaction also cleaves uridine triphosphate (UTP) forming uridine monophosphate and pyrophosphate (PPi). Provide four reasons why UTP can be used to power this reaction (no diagrams necessary).
An experiment was carried out to measure the reaction rate of hydrolysis of acetylcholme (substrate) with serum enzymes (Eadie, 1949). In the experiment, two experiments were conducted, namely experiment 1 without using a prostigmine inhibitor and experiment 2 using a prostigmine inhibitor at 1.5 x 10^-7 mol/l. the data obtained are: a. Is prostigmine competitive or noncompetitive inhibitor? b. determine the value of km and rmax for the two experiments, compare
The phosphorylation and oxidative decarboxylation of oxaloacetate by inorganic phosphate (Pi) to make phosphoenolpyruvate and carbon dioxide is endergonic under intracellular conditions. It is characterized by this equation: Oxaloacetate + Pi ←→ Phosphoenolpyruvate + H2O + CO2 ΔG’ = +24.6 kJ/mol The synthesis of GTP from GDP and inorganic phosphate (Pi) in solution is endergonic under intracellular conditions, and it is characterized by this equation: GDP + Pi ←→ GTP + H2O ΔG’ = +30.5 kJ/mol Write a new net thermodynamically coupled reaction equation that describes the synthesis of phosphoenolpyruvate from oxaloacetate using the hydrolysis of GTP to power the reaction and calculate the new net ΔG’ of this reaction. Show all of your work.
Intramitochondrial ATP concentrations are about 5 mM, and phos- phate concentration is about 10 mM. If ADP is five times more abundant than AMP, calculate the molar concentrations of ADP and AMP at an energy charge of 0.85. Calculate AG for ATP hydrolysis at 37 °C under these conditions. The energy charge is the concentra- tion of ATP plus half the concentration of ADP divided by the total adenine nucleotide concentration: [ATP] + 1/2[ADP] [ATP] + [ADP] + [AMP]
Mutations in adenylate kinase have led to a hyperactive enzyme that ultimately ends up elevating ADP levels in a cell.Calculate the EC (energy charge) given the following atypical adenylate concentrations for the cell containing the mutant adenylate kinase: ATP = 0.5 ??��, ADP = 12.2 ??��, AMP = 80 ??��.

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