I want to use ATR FTIR to accompany the changes in secondary structure of a protein hydrogel by means of deconvoluting the Amide I peak. I gelled my samples and pressed a small amount between two glass slides before analyzing it. This removes some water from the hydrogel. Do you think this could be a correct form for analyzing these hydrogels, or does the removal of water from the hydrogel affect the secondary structure?
P.S: The hydrogels are translucid but closer to opaque (not opaque, but not transparent). They gel through irreversible hydrogen bonds forming beta-sheets mostly.
Though I have no experience in protein analysis, I do have experience with IR and influence of water in the environment and in the sample on the molecular measurement. Dehydrating the sample will most definitely lead to different secondary relationships, maybe try measuring the sample surface with ATR-FTIR carefully, before and after relative times of drying to see the effect on the peaks.
Carolyn Louise Carta I thought so too, but was recommended to do it by a fellow researcher. Thank you for the feedback!
Hi,
I have perfomed some similar analysis. First of all did you work on D2O ? If not it would be impossible to deconvolute your Amide I peak as there is a D2O band in the same region. Moreover if you use D2O, you can follow the amount of water in your gel at 1200 cm-1. I have seen (with HSA, BSA & Myoglobin) that the secondary structure of proteins only change when the band a 1200 cm-1 disapear, but maybe it can depend of the sensibility of your protein.
Good Luck
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