Model proteins genetic engineering

Kiriazis H, Kranias EG 2000 Genetically engineered models with alterations in cardiac membrane calcium-handling proteins. Annu Rev Physiol 62 321—351... 

The extended ternary complex model can take into account the phenomenon of constitutive receptor activity. In genetically engineered systems where receptors can be expressed in high density, Costa and Herz [2] noted that high levels of receptor expression uncovered the existence of a population of spontaneously active receptors and that these receptors produce an elevated basal response in the system. The relevant factor is the ratio of receptors and G-proteins (i.e., elevated levels of receptor cannot yield constitutive activity in the absence of adequate amounts of G-protein, and vice versa). Constitutive activity (due to the [RaG] species) in the absence of ligand ([A] = 0) is expressed as... 

Figure 7.12 The protein engineering cycle. The lower circle represents the genetic engineering activities—the upper the structural determination, modelling and protein characterisation. (See Colour Plate X)...

Genetic engineering. The X-ray structures are known for many hydrolases, allowing for modeling of the substrate in the active site as well as structurally based, random or rational protein mutation to magnify or invert enantioselectivity. An example of the latter is provided by the rational design of a mutant of Candida antarctica lipase (CALB), which, instead of the wild-type R-selectivity, displayed... 

Bacteria produce a wide variety of proteins with toxic effects, and it is possible to detect the potential for such biological activities by sequence homology to known molecules, or by the identification of specific amino acid motifs that are diagnostic for a particular activity (e.g., proteolysis or membranolysis). In these cases, it is often possible to introduce specific mutations by genetic engineering to render the protein nontoxic. Careful selection of the mutations, especially if done in the context of available structural information, such as an atomic-resolution model of the antigen, can produce a fully detoxified molecule that retains the antigenic properties that make it attractive as a vaccine candidate. 

The combination of molecular modeling with genetic engineering to enhance protein stability has been successful in certain cases. For instance, introducing carefully sited novel disulfide bonds increased protein stability in T4 lysozyme (11-13) and in X-repressor (14). However, the results in other proteins, for instance, in subtilisin (15,16) and in dihydrofolate reductase (17) have been less predictable. 

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