I've read some article about collagen extraction from fish waste. Some article said that all procedure should be done at 4-6°C to reduce chain fragmentation. Why it should be done at that temperature, whereas the denaturation temperature of collagen is about 30-40°C. Is it okay if the procedure done under 4°C or more than 6°C?
Thanks
Collagen is usually extracted with pepsin under acidic conditions. So the temperature shold be 4-6oC in case the structure of collagen was damaged, or the activity of pepsin was inhibited.
The so called "extraction" of collagen by pepsin is resulting in fact from the cleavage of the non-triple-helical telopeptides containing the cross-liks on which the insolubility of the collagen fibres is relying. Conversely the resistance to pepsin cleavage of the main structural part of collagen itself is related to the stability of its triple-helical arrangement. Therefore low temperature acts both limiting pepsin activity to the telopeptide cleavage and preventing collagen degradation itself by stabilizing its triple-helical domains. All that is even more important in case of collagen "extraction" from waste material in which collagen is likely to be partly damaged and/or already in conditions to be variably prone to pepsin degradation of its major structural component.
Anyway thank you so much for the answers.
How about doesn't work with pepsin? I mean the extraction only use acid.
If you are doing only acid solubilization of collagen, without pepsin, then the low temperature will decrease the activity of other endogenous proteases in your sample which might damage the collagen, particularly in the non-triple helical domains and/or in less structured domains caused by higher temperatures. Experimentally, try the acid extraction at 4C vs. 20C and see which has a cleaner profile on SDS PAGE.
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