Protein fractionation theory

Although the investigations of both Raunkjaer et al. (1995) and Almeida (1999) showed that removal of COD — measured as a dissolved fraction — took place in aerobic sewers, a total COD removal was more difficult to identify. From a process point of view, it is clear that total COD is a parameter with fundamental limitations, because it does not reflect the transformation of dissolved organic fractions of substrates into particulate biomass. The dissolved organic fractions (i.e., VFAs and part of the carbohydrates and proteins) are, from an analytical point of view and under aerobic conditions, considered to be useful indicators of microbial activity and substrate removal in a sewer. The kinetics of the removal or transformations of these components can, however, not clearly be expressed. Removal of dissolved carbohydrates can be empirically described in terms of 1 -order kinetics, but a conceptual formulation of a theory of the microbial activity in a sewer in this way is not possible. The conclusion is that theoretical limitations and methodological problems are major obstacles for characterization of microbial processes in sewers based on bulk parameters like COD, even when these parameters are determined as specific chemical or physical fractions. 

Fractionation of proteins according to size utilizing cross-linked dextran or polyacrylamide gel columns was first demonstrated by Porath and Flodin 63 in 1959. This technique has become the most widely accepted method for separation and molecular weight determination of hydrophilic and some hydrophobic macromolecules using aqueous buffers with or without organic modifier. While this technique might not be unique in its ability to resolve and separate proteins, it is one additional simple and effective tool in the chemist s armamentarium. The theories behind size-exclusion HPLC and size-exclusion chromatography at low pressure are identical and are described in several publications. 31 34 36 39 44 64 65 ... 

Figure 8.5 Interaction potential for model whey protein layer consisting of densely packed brush-like tethered chains with small a fraction of the whev protein replaced by p-casein chains as represented by a copolymer model. The energy A d) calculated from SCF theory is plotted as a function of surface-surface separation d A, no p-casein B, 2.5% p-casein C, 5% p-casein D, 5% p-casein alone (without whey protein layer). Potentials A, B and D imply that the emulsion system is flocculated potential C implies a stable emulsion state. Reproduced from Dickinson (2006b) with permission.

Transition state theory (Chapter 2, section A) was derived for chemical bonds that obey quantum theory. An equation analogous to that for transition state theory may be derived even more simply for protein folding because classical low energy interactions are involved and we can use the Boltzmann equation to calculate the fraction of molecules in the transition state i.e., = exp(— AG -D/RT), where A G D is the mean difference in energy between the conformations at the saddle point of the reaction and the ground state. Then, if v is a characteristic vibration frequency along the reaction coordinate at the saddle point, and k is a transmission coefficient, then... 

The aims of protein purification, up until the 1940s, were simply academic. To then, even the basic facts of protein structure were not fully appreciated, and pure proteins were needed just to study structure and test the rival theories of the pre-DNA days. During the Second World War, an acute need for blood proteins led to development of the Cohn fractionation procedure for purification of albumin and other proteins from serum (Cohn et al., 1946). This was the inception of large-scale protein purifications for commercial purposes Cohn fractionation continues to be used to this day. 

Lee and Lightfoot [229] developed the theoretical basis of Fl-FFF. This theory has been confirmed by numerous works on the fractionation of model systems, including monodisperse spherical polystyrene latexes and a number of proteins [41,228,229,240], some polydextrans [229], viruses [241], and other spherical particles and macromolecules [242,243]. 

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