Structure-activity correlations mutagenesis

Rosenkranz, H.S. and Klopman, G., Prediction of the carcinogenicity in rodents of chemicals currently being tested by the US National Toxicology Program structure-activity correlations, Mutagenesis, 5, 425 132, 1990. 

Site-directed mutagenesis is generally restricted to the 20 amino acids normally occurring in proteins. Thus, reliance on homologous series of compounds to establish structure-reactivity correlations, a hallmark of mechanistic studies with non-enzymic catalysts, has been lacking with enzymes. One manner in which this limitation can be partially overcome is provided by the demonstration that an enzyme, crippled because of an active-site substitution, can be rehabilitated... 

Cysteine-scanning mutagenesis, involving more than 100 mutations, has been systematically carried out through Cl—C3, the cytoplasmic terminations of TM1-TM7, H8, and the C-terminal tail. In addition, more than 40 pairs of cysteines have been introduced at the cytoplasmic face. With these mutants as a basis set, three classes of experiments have been carried out, namely SDSL, sulfhydryl reactivity, and disulfide cross-linking kinetics. A global comparison of the results provides a unique view of the solution state, its dynamics, and its correlation with the crystal structure. By solution state is meant, in all cases, rhodopsin solubilized in dodecyl maltoside (DM) micelles. The measured functional properties of rhodopsin, namely transducin activation (Resek et al., 1993) and phosphorylation by the rhodopsin kinase (Thurmond et al., 1997), are conserved in this detergent, and it is presumed to be a reasonable approximation to the bilayer environment. 

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