Protein total charge

A method to measure total charge on a protein as it binds to assorted ions is described by M. K. Menon and A. L. Zydney, Measurement of Protein Charge and Ion Binding Using Capillary Electrophoresis, Anal. Chem. 1998, 70, 1581. 

Fig. 6.18. Cubane-type Fe4S4 cluster. The total charge can be +3, +2, or +1 depending on the oxidation states of the iron ions. The cluster shown in the figure is coordinated to four protein cysteines, giving total charge of —1, —2, or —3, respectively.

Proteins bind SDS, and as a first approximation the amount of SDS bound per gram is the same for all proteins. Differences in the amount bound per molecule will result in differences in the total charge, leading to differences in the electrophoretic mobility, and an incorrect value of molecular weight will be inferred. 

Ion-exchange chromatography (IEX-HPLC) is intended to separate proteins based on differences in their total charge. Both anionic and cationic exchange modes are available. This technique can potentially detect and... 

A.26.4 The problem states that charge is related to the number of amino acids and those can be estimated assuming 0.1 kD = 1 amino acid, and we can assume 2 amino acids equal 1 SDS molecule = one negative charge. The total charge added to each protein is then it s molar mass divided by 0.2 kD. (a) 337.5, (b) 62, (c) 1650, (d) 340, (e) 90. 

At or in the proximity of their pi values proteins exhibit a minimum total charge and reduced solubility in the electrolyte solution [350,370,385]. This increases the probability of aggregation, and is further enhanced by the low ionic strength of the ampholyte buffer. Under these conditions protein precipitation results from hydrophobic interactions. These interactions can be suppressed by addition of additives such as ethylene glycol (10-40 %), non-ionic or zwitterionic surfactants (1-4 %), or sorbitol to the ampholyte buffer. 

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