Open AccessHydrogen‐Ion‐Titration Studies of Pancreatic Phospholipase A and Its Zymogen
Author(s) -
Janssen Lambert H. M.,
Bruin Simon H.,
Haas Gerard H.
Publication year - 1972
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1972.tb01897.x
Subject(s) - titration curve , titration , chemistry , potentiometric titration , ionic strength , titratable acid , acid–base titration , inorganic chemistry , ion , biochemistry , organic chemistry , aqueous solution
The titration curves of porcine pancreatic phospholipase A an its zymogen were measured in the pH region 2.0 to 9.5 at 25°C. In the pH region 2 to 6 the titration curves were measured using an Ag‐AgCl electrode as reference electrode, at ionic strength 0.5. It was found that the titration of the carboxyl groups could be described with the Linderstrøm‐Lang model using an intrinsic pK of 4.5 and an electrostatic interaction factor ω of 0.073. The side‐chain carboxyl group of the second ( viz. glutamyl) residue in prophospholipase appeared to have an abnormally low pK of 3.7. The maximum positive charge of the two proteins as determined by titration agreed well with the values predicted by the amino acid composition. From an analysis of the titration curves covering the region 4.5 to 9.5, which were measured with a calomel electrode at ionic strength 0.1, the following could be concluded. The intrinsic pK and ω of the carboxyl groups appeared to be at this ionic strength 4.7 and 0.114, respectively. In both phospholipase and prophospholipase the number of titratable carboxyl groups was found to be 3 higher than given by the amino acid analysis. All 3 histidines present were titratable with an intrinsic pK of 6.6 and ω near zero. The terminal amino group which is free only in phospholipase appeared to have an apparent pK of 8.3.