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Calculation of the isoelectric points of native proteins with spreading of p K a values
Author(s) -
Henriksson Gunnar,
Englund AnnKristin,
Johansson Gunnar,
Lundahl Per
Publication year - 1995
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.11501601227
Subject(s) - chemistry , isoelectric point , isoelectric focusing , sialic acid , amino acid , side chain , residue (chemistry) , urea , immobilized ph gradient , chromatography , crystallography , biochemistry , organic chemistry , enzyme , polymer
The isoelectric points (p I ) of native proteins are important in several separation techniques. For estimating p I values the net charge of several proteins was calculated versus pH by use of the Henderson‐Hasselbalch equation. Amino acid composition, p K a values for amino acid side chains and for the N ‐ and C ‐terminal groups, and the presence of other charged groups were taken into account. A set of p K a values was chosen for amino acid residues with ionizable side chains. Each particular type of ionizable group was assumed to have p K a values distributed around the chosen value, thereby simulating the situation in proteins and polypeptides. The calculated p I values showed reasonably good agreement with experimental ones for most of 16 native proteins over a wide pH range (3.4–11) when charge contributions of heme groups, sialic acid residues, etc ., were taken into account. The calculated p I for the human red cell glucose transporter (Glut 1) with one sialic acid residue was decreased from 8.8 to 8.5 by introducing p K a value spreading and became consistent with the experimental p I value of 8.4 ± 0.05 at 15°C determined in the presence of 6 M urea. The p I of the native Glut1 was lower, 8.0 ± 0.1, at 22°C. In general, the p I values for native proteins are affected by the three‐dimensional structure of the proteins, which causes greater differences between calculated and experimental p I values than in the case of polypeptides for which p I values are determined in the presence of urea.