Tagged Questions

Enzyme kinetics are the studies of the biophysical properties of enzymes and their substrates.

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2
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2answers
63 views

Why is Eadie- Hofstee Plot not linear

Besides the reason of cooperativity between multiple active sites on an enzyme, what are the other reasons for the Eadie-Hofstee plot to be non-linear
-2
votes
0answers
36 views

Eadie-Hofstee Plot [on hold]

If an eadie-hofstee plot shows two distinct gradient, Does this imply cooperativity between the two active sites? and how do we find the hill coefficient? The question asked us to plot graphs of V ...
-1
votes
0answers
30 views

Is autocatalysis a non-Michaelis-Menten Reaction [on hold]

If a reaction involves its own product altering the rate of reaction and the affinity to the substrate, does it still obey the Michaelis-Menten kinetic model?
1
vote
0answers
24 views

Enzyme Assay - pectinase

During assaying an enzyme at high temperature, the substrate (Pectin) is degraded by the high temperature rather than by enzyme, so, how can I minimize degradation of the substrate by the temperature? ...
1
vote
1answer
28 views

can a bacterial protease inhibitor cocktail be used for Western Blotting involving HUVEC cells and HT-29 adenocarcinoma cells?

Dear fellow biochemists, I need some advice on Western Blotting, more specifically the use of certain protease inhibitors with the RIPA cell lysis buffer and protease inhibitor cocktail. A Millipore ...
2
votes
1answer
54 views

Pectinase Enzyme Assay

I am working on pectinase enzyme assay. I incubated 900 ul of substrate for 10 minutes in the water bath, followed by adding 2ml of DNSA reagent, then 100ul of enzyme extract added finally i read the ...
4
votes
1answer
88 views

DpnI over-digestion

We have a long protocol that we are optimizing that includes DpnI digestion of a PCR product (to remove any of the template DNA if it's methylated, and while we're not certain in the blind tests, ...
2
votes
1answer
49 views

What does “thermodynamic equilibrium” mean for an enzyme-substrate complex?

From Fersht, Enzyme Structure and Mechanism p. 87: The Michaelis-Menten mechanism assumes that the enzyme-substrate complex is in thermodynamic equilibrium with free enzyme and substrate. In ...
1
vote
0answers
114 views

What's the difference between Ki and IC50?

I have just read this post at researchgate: http://www.researchgate.net/post/How_to_calculate_experimental_binding_free_energy_from_the_IC50_value I would not be very sure about what the difference ...
1
vote
1answer
44 views

Enzyme kinetics [closed]

I can't understand how to study enzyme kinetics. Say I have a lipase and want to study the kinetics of this lipase using a fluorogenic substrate, how would I do this? From what I understand I would ...
2
votes
1answer
35 views

Activity of glucokinase

From Solomon et al, 2013 ACC Synthetic biology and from this video : Here, there are 2 competing reactions for glucose - one with glk as enzyme and other with gdh as enzyme. In the graph, y axis ...
3
votes
3answers
123 views

Are there any methods to quantify H2O2 (hydrogen peroxide) which don't rely on horseradish peroxidase?

I can't use HRP since my substrates interfere with it and I need a real-time method because I want to measure kinetic parameters of some oxidases.
4
votes
2answers
349 views

Why are enzyme-catalysed reactions slower at lower substrate concentration?

Suppose I'm using 200 nmoles of enzyme and 2 mmoles of substrate. The enzyme should be saturated but if I use 50 mmoles of substrate, the reaction will be faster. Why? I just can't get it! Even at ...
1
vote
1answer
90 views

Modeling yeast biochemical pathway using enzyme kinetics

I need enzyme concentration and metabolites concentration values to en-corporate these values in my model and do some simulation. I searched through some database yeast metabolome database ...
1
vote
1answer
148 views

What is the use of futile reaction cycling such as Fruc-P to Fruc-BP?

One rate-limiting step of glycolysis is the conversion of Fructose-Phosphate (Fruc-P) to Fructose-Bisphosphate (Fruc-BP), catalysed by Phosphofructokinase (PFK). The reaction involves hydrolysing one ...