Protein alternative approaches

Including the solvent around a protein can be done, in principle, by the explicit approach of Chapter 3. Such a treatment, however, is very expensive, in view of the large number of water molecules needed to properly solvate the entire protein. Thus we will consider below two alternative approaches which allow one to effectively represent the solvent. The discussion of these models will be focused on electrostatic aspects where the importance of solvent effects is easily demonstrated. 

An alternative approach is to synthesize an artificial gene in the test-tube starting with the appropriate deoxyribonucleotides. This approach, which demands that the entire amino acid sequence be known, has been used to clone genes encoding proteins 200 amino acids long. 

An alternate approach for biomolecule recover is to employ degradable surfactants [157]. A series of nonionic surfactants has been synthesized that contain the acidic pH-degradable cyclic ketal linkage [153]. These surfactants readily form w/o-MEs when at neutral pH or higher but, the surfactants readily degrade at moderately low pH (ca. 5), releasing the encapsulated aqueous phase and its constituents. Work is ongoing to develop these surfactants in w/o-ME protein extraction processes [153]. 

In the above sense, evolution cannot be described by reference to molecules much though speciation may be referenced by DNA (RNA) or protein content. Speciation may be only a way to characterise evolution and not the way in which to understand it. The global evolution of an ecosystem of living chemistry is an alternative approach. The parameters which can evolve and did evolve are as follows ... 

An alternative approach is that adopted by Horrocks and co-workers, where the aromatic residues in metal-binding proteins are used as sensitizers. Since the distance between the metal and the donor is effectively fixed, this provides a rigid scaffold for the experiment, and the absence of a directly conjugated pathway between the metals means that Forster (through space) energy transfer can be assumed. The r-6 distance-dependence of this means that the extent of sensitized emission from the lanthanide ion provides information on the spatial relationship between the metal-ion binding site (lanthanide ions often bind at Ca2+ sites) and nearby aromatic residues. 58-60... 

A-chain immunotoxins, however, may not be quite as cytotoxic as conjugates formed from intact toxin molecules (Manske et al., 1989). In an alternative approach to A chain use, the intact toxin of two-subunit proteins is directly conjugated to a monoclonal without isolation of the A chain. Conjugation of an antibody with intact A-B chain toxins can be done without a cleavable linker, as long as the A chain can still separate from the B chain once it is internalized. Therefore, it is important to avoid intramolecular crosslinking during the conjugation process which can prevent release of the A-B complex. In addition, since the B chain... 

A number of approaches may be adopted in an attempt to reduce or eliminate protein im-munogenicity. Protein engineering (Chapter 3), for example, has been employed to humanize monoclonal antibodies (Chapter 13). An alternative approach entails the covalent attachment of polyethylene glycol (PEG) to the protein backbone. This can potentially shield immunogenic epitopes upon the protein from the immune system. 

An alternative approach to clone the receptor was taken by Thomas and colleagues (1990, 1991). They screened a cDNA library from peritoneal rabbit cells with an antisense oligonucleotide probe deduced from the coding region of the second transmembrane domain, which is common to G-protein-coupled receptors (fMet-Leu-Phe is a G-protein-linked receptor). Screening of this library yielded a putative receptor clone F3R, characterised as follows ... 

An alternative approach to obtaining a starting model for the refinement that does not need NOE data is called molecular fragment replacement [71] and treats the protein of interest as overlapping fragments of 7 to 10 residues in length. A protein database is... 

Use of zwitterions is an alternative approach that provides more effective mobilization of protein zones across a wide pH gradient.83 For example, cathodic mobilization with a low-pi zwitterion enables efficient mobilization of proteins with pis ranging from 4.65 to 9.60. The proposed mechanism for zwitterion mobilization couples a pH shift at the proximal end of the tube with a displacement effect at the distal end as the zwitterion forms an expanding zone within the gradient at its pi. Effective zwitterion mobilization depends on the selection of the appropriate mobilization reagent. 

Nagai and Thergersen found that an alternative approach was required to achieve efficient expression of the -chain subunits of human hemoglobin in E. coli [55]. In this case, P-globin was produced as a fusion protein that possessed the sequence Ile-Glu-Gly-Arg between the 31 amino-terminal residues of X cll... 

The alternative approach is to use the knowledge gained about the enzymes, to construct chemical catalysts. Here the field is in its infancy, since the construction of structural models could only start when the structure of the proteins began to emerge, in 1995. Previously, many heterogeneous catalysts had been known, for example for hydrogenation processes. But most of these used heavier, and scarcer metals. 

An alternative approach is to use antibody fragments (the Fab domian) raised against a particular protein. This technique has been used with some success for crystallizing a number of membrane proteins, including the CIC chloride channel (Dutzler et al., 2002), a K+ voltage-dependent channel (Jiang et al., 2003), and a cytochrome c oxidase (Iwata et al., 1995). 

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