About the protein enginerring - In urgent, thanks (Apr/15/2007 )

If i get the enzyme X, it was known that the tyrosine residue near the active site, how can i do to confirm the importance of this residue.

Thanks

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two suggestions:

a functional Y may be relevant in a phosphorylated/dephosphorylated state

1. more accepted but some work is mutagenesis of Y to D (phosphomimetic) as well as Y to A (non-phosphorylatable);


2. if there is a phospho-specific Ab for this site available you can demonstrate dependence of phospho-/non-phospho Y with the function of the enzyme; if it is the only Y in the protein (what would rarely be the case) you can also use a global anti-phospho-Y Ab

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