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The Importance of Correct Protein Concentration for Kinetics and Affinity Determination in Structure-function Analysis

DOI :

10.3791/1746-v

19:16 min

March 17th, 2010

March 17th, 2010

20,110 Views

1GE Healthcare Bio-Sciences AB

We apply label-free protein interaction analysis using Biacore X100 for structure-function analysis of the binding of several cystatin B mutants to papain through kinetic characterization. Calibration-free concentration analysis (CFCA) measures the concentration of protein with retained binding activity without the need for a standard curve. We show that confirmation of concentrations using CFCA increases the reliability of the kinetic analysis and that kinetic constants can reliably be determined even if the activity of a recombinant protein is reduced.

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Protein Concentration

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