Molecular interaction between apo or holo alpha-lactalbumin and lysozyme: formation of heterodimers as assessed by fluorescence measurements.
In a previous work, we reported that contrary to native calcium-loaded alpha-lactalbumin (holo alpha-LA), calcium-depleted form (apo alpha-LA) has the ability to self-assemble with lysozyme (LYS) to form different supramolecular structures in temperature-dependent manner. In this study, we examine what happens at molecular scale using fluorescence techniques. Fluorescence anisotropy coupled with fluorescence lifetime measurements provides a means to measure intermolecular interactions. We showed that LYS interacts with both apo alpha-LA and holo alpha-LA to form oligomers, assumed to be heterodimers, at 10 degrees C and 45 degrees C. The dissociation constants for dimerization were found to be in the muM range and increased significantly with increasing ionic strength from 39 to 124 mM. Although the binding constants of holo alpha-LA-LYS and apo alpha-LA-LYS complexes were of the same order of magnitude, the shape or conformation of formed heterodimers differed as assessed by fluorescence parameters in particular correlation time calculations. Such conformation differences could explain why holo alpha-LA-LYS complexes are trapped as heterodimers while the apo alpha-LA-LYS complexes have the ability to further self-assemble into various supramolecular structures.
Nigen
Michaël
M
INRA-agrocampus Rennes, UMR 1253, STLO, 65 rue de Saint-Brieuc, F-35042, Rennes, France.
Le Tilly
Véronique
V
Croguennec
Thomas
T
Drouin-Kucma
Delphine
D
Bouhallab
Saïd
S
D000818:Animals; D001059:Apoproteins / Q000737:chemistry* / Q000378:metabolism; D002417:Cattle; D002645:Chickens; D003619:Dansyl Compounds / Q000378:metabolism; D005454:Fluorescence Polarization; D007700:Kinetics; D007768:Lactalbumin / Q000737:chemistry* / Q000378:metabolism; D009113:Muramidase / Q000737:chemistry* / Q000378:metabolism; D009994:Osmolar Concentration; D011485:Protein Binding; D011487:Protein Conformation; D055503:Protein Multimerization; D013050:Spectrometry, Fluorescence; D013696:Temperature
D001059:Apoproteins; D003619:Dansyl Compounds; D007768:Lactalbumin; C421864:hen egg lysozyme; D009113:Muramidase; C029038:dansyl chloride