By detecting the Km and Vmax values of the crude and pure enzyme, it was found that the Vmax of the pure enzyme remained the same, but the Km value increased.
That is, the Km of the pure enzyme was larger than that of the crude enzyme, which indicated that the pure enzyme had competitive inhibitors, while pure enzyme didn't contain any inhibitor, so it didn't conform to the principle.
Is there any difference between crude enzyme and pure enzyme in catalytic behavior?
Because of their different specificities (specific contents) for the same substrate.
The first thing to consider is whether the crude enzyme contained only a single enzyme with the measured activity. Could there have been multiple enzymes catalyzing the same reaction?
'According to Michaelis - Menten equation, Km is equal to substrate concentration at which the velocity is half the maximum Vmax.
In competative inhibition Vmax is always constant, but Km is alter (increased) showing the substrate specificity to enzymes.
When learning the catalytic activity of enzymes pure enzymes always shows more Km.
Pure enzyme is the only monomeric enzyme, but crude enzyme is oligomeric
So firstly confirm the crude enzyme (oligomeric) and then compare the Km between pure and crude enzyme.
Was that under the same conditions like freshly made buffers, concentration, pH & incubation time and temperature for both (pure & crude) Are you using the right substrate for the enzyme? sometimes many enzymes in crude extract degrade the same substrate at the same time and we linked that to the enzyme of our interest which is wrong. if your enzyme is pure and you are very certain of using this substrate as the best for this enzyme then you may need to rethink about buffer, pH, incubation temperature & time. finally bear in mind that the activity of an enzyme decreases if stored for longer time. if you are not sure of how to calculates Km then see the following link
https://www.sciencedirect.com/topics/immunology-and-microbiology/michaelis-constant#:~:text=TEMPERATURE%20%7C%20Proteins%20and%20Temperature&text=Thus%2C%20Km%20has%20units,reach%20a%20certain%20reaction%20rate).
Hope this helps
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