for CAT activity, is there is any threshold regarding the concentration of protein in the enzyme has to be taken into consideration before doing the experiment.
You have to standarize the concentration. You will have to run different reactions using different concentration of the protein. From those reactions, you will have to select one concentration.
Hi there,
It should be catalytic concentration! Which means the substrate should be in a large excess versus the enzyme (at least 2 orders of magnitude). Apart from this the conditions have to be defined empirically according to the actual catalytic capacity of your enzyme sample and to the technical constraints linked to the method of detection. What you are actually looking for is a condition for which the reaction occurs at constant rate for a sufficient laps of time (for initial velocity determination).
Biotechniques. 1994 Jul;17(1):36, 38, 40.
You may check this article.
Please check
1. Olson BJ1, Markwell J. Assays for determination of protein concentration. Curr Protoc Protein Sci. 2007 May;Chapter 3:Unit 3.4. doi: 10.1002/0471140864.ps0304s48.
2. He, F. (2011). Bradford Protein Assay. Bio-101: e45. DOI: 10.21769/BioProtoc.45.
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