I need to study the interaction of my compound with BSA protein using simple UV, Fluorescence and IR techniques. Once I prepare the protein solution in buffer, how should I confirm that the protein is in the native form before beginning any experiment?
The following link may be useful.
https://hal.inria.fr/hal-00895648/document
http://pubs.acs.org/doi/pdf/10.1021/ac00279a044
https://www.london-nano.com/research-and-facilities/highlight/detecting-protein-denaturation-and-the-interactions-between-protei
http://physinfo.uark.edu/largeimages/JeffSparksHonorsThesis.pdf
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5355583/pdf/kosfa-37-44.pdf
Hi there,
UV spec, light scattering, circular dichroism should tell you.
You can perform an native PAGE (without SDS) and a SDS-PAGE in order to see if there are different bands. However, with an UV-vis spectrum is usually enough to know if your protein is not denatured.
But what are the changes I will observe in the UV spectrum if at all the protein has denatured?
If you do not see typical UV-VIS spectrum (see attached image) with a maximum peak at 280 nm (or between 278-281 nm) it seems that the protein could be denatured. In addition, you have to check the activity and compare with activity values reported in literature
Hi there,
UV spectra may be informative only on the presence of aggregates in the sample. If no aggregation, spectra should come back to zero between 320 and 340nm , if presence of aggregates the baseline between 320 and 340 would be higher due to light diffusion by aggregates.
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