Characterization of human and bovine phosphatidylethanolamine-binding protein (PEBP/RKIP) interactions with morphine and morphine-glucuronides determined by noncovalent mass spectrometry
Cedric Atmanene, Alexis Laux, Elise Glattard, Arnaud Muller, Francoise Schoentgen, Marie-Helene Metz-Boutigue, Dominique Aunis, Alain Van Dorsselaer, George B. Stefano, Sarah Sanglier-Cianferani, Yannick Goumon
Med Sci Monit 2009; 15(7): BR178-187
The phosphatidylethanolamine-binding protein (PEBP/RKIP), initially found to bind phosphatidylethanolamine (PE), has been shown to be associated with morphine derivatives. Our recent study on bovine primary chromaffin cells showed that inside secretory granules, PEBP is noncovalently associated to endogenous morphine-6-glucuronide (M6G), a highly analgesic morphine metabolite. During stress, M6G-PEBP complexes may be released into circulation to target peripheral opioid receptors. We now report the investigation of PEBP binding properties towards morphine and morphine analogs.
Material and Method: Noncovalent electrospray ionization mass spectrometry (ESI-MS) was used to investigate bovine and human PEBP binding properties towards morphine and morphine-glucuronides.
Results: We describe for the first time that: (i) PEBP directly interacts with morphine glucuronides (M3G and M6G) but not with morphine, (ii) that the presence of a glucuronide group either on the 3rd or the 6th morphine's carbon does not affect these interactions, (iii) that M6G binds PEBP in a similar manner as the reference ligand PE and (iv) that PEBP displays a similar affinity for PE, M6G and M3G.
Conclusions: Our results suggest that PEBP might protect M6G following its secretion into blood, leading to a longer half life. This study highlights the potentialities of ESI-MS to validate / invalidate the formation of protein: ligand noncovalent complexes when low affinity binders (i.e., compounds with affinities lower than 10(3) M(-1)) are concerned.
Keywords: Phosphatidylethanolamines - metabolism, Phosphatidylethanolamine Binding Protein - metabolism, Morphine Derivatives - metabolism, Mass Spectrometry, Humans, Cattle, Animals, Titrimetry