Extracting alpha amylase from human saliva

[Tiberius]

I wanted to perform some experiments on enzyme rates and chose amylase as it is easy to obtain. However, I wanted it in pure form so that I can perform mass/concentration-based calculations as well.

Is there any way to extract alpha-amylase from human saliva?

 

Thank you.

[ewmon]

Is there any way to extract alpha-amylase from human saliva?

Unless you have access to a chromatography column, it'll probably need to be precipitated out, so I googled alpha amylase precipitation and found

 

PURIFICATION OF SALIVARY α-AMYLASE

Edited November 4, 2010 by ewmon

[Tiberius]

Thanks for your reply.

I may get access to column chromatography facilities in a BSc Chem Lab... Assuming I do, could you point me to a link for the method ?

 

Also, do you have any experience with crystallizing out(and growing) thermally unstable proteins like alpha amylase ?

 

Thank you.

[ewmon]

Sorry, I have yet to use LC and have done very little precipitating of proteins.

[CharonY]

Crystallization is tricky. I would go for a crystallization paper and try that protocol. However, generally you would need to purify it in relatively high concentration first (and with high purity), which is non-trivial, if using body fluids. In saliva you will find quite a number of different proteins. LC separation could work, though a single separation (e.g. reverse-phase) may be insufficient to get a pure fraction.

 

Other possibilities include affinity purification, but most would rather use an overexpression system and purify it from there for crystallography studies.