Title: A ca2+-binding chimera of human lysozyme and bovine alpha-lactalbumin that can form a molten globule
Authors: Pardon, E
Haezebrouck, Petra
Debaetselier, A
Hooke, Sd
Fancourt, Kt
Desmet, Johan
Dobson, Cm
Van Dael, Herman
Joniau, Marcel #
Issue Date: May-1995
Publisher: Amer soc biochemistry molecular biology inc
Series Title: Journal of Biological Chemistry vol:270 issue:18 pages:10514-10524
Abstract: In contrast to lysozymes, which undergo two-state thermal denaturation, the Ca2+-free form of the homologous alpha-lactalbumins forms an intermediate ''molten globule'' state, To understand this difference, we have produced a chimera of human lysozyme and bovine alpha-lactalbumin, In the synthetic gene of the former the sequence coding for amino acid residues 76-102 was replaced by that for bovine alpha-lactalbumin 72-97, which represents the Ca2+-binding loop and the central helix C, The chimeric protein, LYLA1, expressed in Saccharomyces cerevisiae was homogeneous on electrophoresis and mass spectrometry, Its Ca2+ binding constant was 2.50 (+/-0.04) x 10(8) M(-1), and its muramidase activity 10% of that of human lysozyme, One dimensional NMR spectroscopy indicated the presence of a compact, well structured protein, From two-dimensional NMR spectra, main chain resonances for 118 of a total of 129 residues could be readily assigned, Nuclear Overhauser effect analysis and hydrogen-deuterium exchange measurements indicated the presence and persistence of all expected secondary structure elements, Thermal denaturation, measured by circular dichroism, showed a single transition temperature for the Ca2+ form at 90 degrees C, whereas unfolding of the apo form occurred at 73 degrees C in the near-UV and 81 degrees C in the far-UV range. These observations illustrate that by transplanting the central part of bovine alpha-lactalbumin, we have introduced into human lysozyme two important properties of alpha-lactalbumins, i.e. stabilization through Ca2+ binding and molten globule behavior.
ISSN: 0021-9258
Publication status: published
KU Leuven publication type: IT
Appears in Collections:Faculty of Science, Campus Kulak Kortrijk
Electrical Engineering - miscellaneous
Faculty of Medicine, Campus Kulak Kortrijk
# (joint) last author

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