Stabilization of lysozyme by the introduction of Gly-Pro sequence.
Three mutant lysozymes where the Asp101-Gly102 sequence of lysozyme was converted to Asp101-Pro102, Gly101-Pro102 and Pro101-Gly102 were prepared to investigate the effect of proline residues on the stabilization of proteins. The free energy changes of lysozymes for the unfolding in aqueous solution at pH 5.5 and 35 degrees C were 10.0, 10.1, 11.0 and 7.7 kcal/mol for wild type, Asp101Pro102, Gly101Pro102 and Pro101Gly102 lysozyme respectively. When the energy level in the unfolded state of wild type lysozyme was fixed at a standard level, the energy levels in the folded state of Asp101Pro102 and Pro101Gly102 lysozymes were found to be higher than that of wild type lysozyme on the basis of delta GD(H2O) and entropy losses of their polypeptide chains in the unfolded state. The presence of some strain in the folded state of these lysozymes was supported by both the calculation of conformational energy for a trans-L-prolyl residue [Schimmel, P.R. and Flory, P.J. (1968) J. Mol. Biol., 34, 105-120] and the analysis of structures of energy-minimized mutant lysozymes. Therefore, it is concluded that the formation of the Gly-Pro sequence is effective in avoiding possible strain in the folded state of a protein caused by the introduction of proline residue(s).
Ueda
T
T
Faculty of Pharmaceutical Sciences of Kyushu University, Fukuoka, Japan.
Tamura
T
T
Maeda
Y
Y
Hashimoto
Y
Y
Miki
T
T
Yamada
H
H
Imoto
T
T
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002942:Circular Dichroism; D006358:Hot Temperature; D008969:Molecular Sequence Data; D009113:Muramidase / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D011392:Proline; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000378:metabolism
D011993:Recombinant Fusion Proteins; D011392:Proline; D009113:Muramidase