Unnatural amino acid and related methods provided a special mechanism to implement isotope incorporation in a specific site of a specific membrane protein.The site specific incorporation resulted in a single peak in NMR spectrum,resulting in unambiguous resonance assignment.The protein specific labeling makes the in situ membrane protein analysis using solid state NMR.With substitute unnatural amino acids of 3,5-difuoro-tyrosine(F2Y)with tyrosine,tyrosine phosphorylation analysis of the essential auto-phosphorylaiton site of an integral tyrosine kinase in E.coli membrane was conducted and the data indicate a quick dephosphorylation even in the presence of excessive ATP and Mg2+.The same methods have also been applied for structure and function analysis of transporters.