Salt concentration, association process

Hydrophobic interaction chromatograph (HIC), while very attractive in principle, has proved difficult to scale up for processing. A recent series of articles explores some of the unique problems associated with process-scale HIC. Load sample preparation20 must be carefully examined to prevent protein aggregate formation in the presence of the relatively high salt concentrations used in this technique. Successful scale-up also requires the setting of wide specifications to accomodate routine variations in the feed.21 The effect of the salt concentration on capacity may be somewhat more... 

Kinesin and dynein are two microtubule-associated ATP-dependent motors responsible for intracellular motility. Gilbert et al made direct measurements of the dissociation kinetics of kinesin from microtubules (MTs), the release of orthophosphate and ADP, and the rebinding of this motor to MTs. They observed processivity in ATP hydrolysis amounting to 10 molecules ATP per site at low salt concentration and 1 molecule of ATP per site at a higher concentration of salt. After hydrolysis, the dissociation of kinesin from the MT is rate-limiting, and rebinding of kinesin-ADP to MTs is fast. The authors discuss how this behavior differs from that of skeletal myosin. 

The amount of antibody initially bound to antigen will be determined by the association rate and incubation time, and of course by the level of antigen expression. The association rate is influenced by the association rate constant, antibody concentration, and temperature. Other factors, such as salt concentration and pH, are also important, but these are usually controlled within physiological limits. Further processes before analysis (washing, fixation, storage) may result in loss of some of the antibody initially bound the amount lost will depend on the dissociation rate and valency, and will also be influenced by temperature. 

Association reactions can be characterized by equilibrium constants. Experimental determination of equilibrium constants for each step in an association reaction provides vital information about the properties of the associating system. In particular, the mode of association (e.g., monomer-dimer, monomer-tetramer, indefinite), and the strength of the association (that is, the degree to which various oligomers can exist at various total concentrations) can be obtained. The evaluation of equilibrium constants over a range of solution conditions (such as salt concentration and temperature) can be used to obtain information on the enthalpy and entropy of the various steps in the association and the types of bonds involved in the assembly process. Note that this information can be obtained in the complete absence of structural information, although, of course, any available structural information can be used to aid in the interpretation of the thermodynamic data. 

Cmp and Dmp are specific constants related to the dimensions of the macromolecule, while the coefficient Amp is inversely dependent on protein size. A similar relationship can be derived for the stationary phase component of the change in the Gibbs free energy due to electrostatic effects (AG >sp><), although the precise relationship between salt concentration in the mobile phase and activity coefficient of the protein when bound to the sorbent may take a more complex form. The electrostatic free-energy change associated with the chromatographic retention process then can be expressed as... 

Analyte ion association (pH and salt concentration-dependent process)... 

The dynamics of the evolution of the blue shift and of the association process exhibit linear dependence on the overall concentration of the salt rather than on the concentration of the free ions (6), and in the case of TBABr yield a pseudo-unimolecular rate of 7.7 X 10 M" s . This finding is not surprising considering the low dissociation constants of salts in XHF (from -1 X 10" to 1 X 10" ). Even at a 1 mM concentration, the salts are less than 10% disso-... 

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