Stabilization of a protein by removal of unfavorable abnormal pKa: substitution of undissociable residue for glutamic acid-35 in chicken lysozyme.


Abstract

Glu35 in chicken lysozyme has an abnormally high pKa (6.1) partly due to the hydrophobic environment provided by Trp108. The relationship between protein stability and abnormal pKa was investigated in detail by using mutant lysozymes in which Glu35 was replaced by undissociable residues and an oppositely ionizable residue. It was found that lysozyme was stabilized at alkaline pH range by the replacement of Glu35 with an undissociable residue, Gln (E35Q lysozyme) or Al (E35A lysozyme). On the other hand, when Glu35 was replaced by His (E35H lysozyme), which could have an opposite charge to Glu by ionization, the introduced His35 was found to have an abnormally low pKa (3.6), leading to the destabilization of lysozyme at acidic pH. These observations are completely consistent with the situation that the environment around Glu35 is highly hydrophobic and therefore the placement of either a positive or negative charge in such an environment leads to destabilization of lysozyme. These observations also indicate that the replacement of an acidic residue having abnormally high pKa or a basic residue having abnormally low pKa by an undissociable residue is a very efficient and general method for stabilization of a protein. Study holds ProTherm entries: 1749, 1750, 1751, 1752, 1753, 1754, 1755, 1756, 1757, 1758, 1759, 1760, 1761, 1762, 1763, 1764, 1765, 1766, 1767, 1768 Extra Details: chicken lysozyme; glutamic acid; protein stability;,hydrophobic; stabilization

Submission Details

ID: 3DfW5ByY3

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:17 p.m.

Version: 1

Publication Details
Inoue M;Yamada H;Hashimoto Y;Yasukochi T;Hamaguchi K;Miki T;Horiuchi T;Imoto T,Biochemistry (1992) Stabilization of a protein by removal of unfavorable abnormal pKa: substitution of undissociable residue for glutamic acid-35 in chicken lysozyme. PMID:1390669
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