Isolation and characterization of 101-succinimide lysozyme that possesses the cyclic imide at Asp101-Gly102.
Lytic activity of lysozyme solution gradually increased on incubation at pH 4, 40 degrees C. When the solution was analyzed by use of cation-exchange HPLC at pH 5, a new peak appeared with increased incubation time. The derivative in the new peak was identified to be 101-succinimide lysozyme in which cyclic imide formed at Asp101-Gly102. The formation of 101-succinimide lysozyme increased with increases in concentration of acetate buffer. Kinetic analysis of the formation of 101-succinimide lysozyme indicated that the cyclic imide was stable below pH 5 due to suppression of the hydrolysis of cyclic imide. Its lytic activity against M. luteus, which has a negative charge, was 165% at pH 7, whereas its activity against glycol chitin, which has no charge, was 90%. Since the lytic activity of Asn101 lysozyme, where one negative charge is eliminated, reached a maximum of 125%, it was suggested that the increase of lytic activity against bacterial cells in 101-succinimide lysozyme was due not only to the disappearance of the negative charge at Asp101 but also to the removal of steric hindrance at the upper part of the active site cleft.
Graduate School of Pharmaceutical Sciences, Kyushu University 62, Fukuoka, Japan.
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Tomizawa
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Yamada
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Ueda
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Imoto
D000085:Acetates / Q000737:chemistry; D000818:Animals; D002021:Buffers; D002645:Chickens; D002686:Chitin / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D009113:Muramidase / Q000737:chemistry*; D011485:Protein Binding; D013329:Structure-Activity Relationship; D013388:Succinimides
D000085:Acetates; D002021:Buffers; D013388:Succinimides; D002686:Chitin; D009113:Muramidase