Enzyme mutation studies

Are mutation studies necessary Enzyme studies or other phenotypic or functional investigations may be more sensitive and more (cost-)effective for reaching a diagnosis. 

Even when an enzyme is studied exhaustively, identifying the amino acid residues that control the catalytic activity is a difficult task In fact, site-specific single mutations rarely do their job, since single mutations far from the active site can change the protein s properties. 

Unno, H., ichimaida, R, Suzuki, H., Takahashi, S., Tanaka, Y, Saito, A., Nishino, T., Kusunoki, M. and Nakayama, T. (2007) Structural and mutational studies of anthocyanin malonyltransferases establish the features of BAHD enzyme catalysis. /. Biol. Chem., 282, 15812-22. 

Sebastian S, Wilson JE, Muhchak A, Garavito RM. Allosteric regulation of type I hexokinase A site-directed mutational study indicating location of the functional glucose 6-phosphate binding site in the N-terminal half of the enzyme. Arch Biochem Biophys 1999 362 203-10. 

B shows a result obtained for the E. coli enzyme, and studies with mutations that shift the [4Fe-4S] reduction potential have shown that the boost in catalytic rate correlates closely with reduction of this relay centre. ... 

As mentioned above, the quinazoline aspartates are potent inhibitors of DHFR, and in fact, may be more tightly bound to the enzyme than either MTX or AMT. Albrecht et al. [268] observed essentially stoichiometric inhibition of DHFR s from two lines of Chinese hamster cells, and also noted that, depending on the enzyme studied, dihydrofolate was much less successful in reversing the inhibitory effect of methasquin than of MTX or AMT. This was especially true when the dose-response for inhibition of enzyme activity by MTX and AMT displayed a curvilinear pattern indicative of a relatively high off-rate . Since this property was perceived as a likely basis for acquired antifolate resistance via enzyme mutation, the quinazoline aspartates were seen as possessing a distinct advantage over MTX and AMT. 

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