title: How High Concentrations of Proteins Stabilize the Amorphous State of Calcium Orthophosphate: A Solid-State Nuclear Magnetic Resonance (NMR) Study of the Casein Case.
abstract: Understanding how proteins stabilize amorphous calcium ortho-phosphate (ACP) phases is of great importance in biology and for pharmaceutical or food applications. Until now, most of the former investigations about ACP-protein stability and equilibrium were performed under conditions where ACP colloidal nanoclusters are surrounded by low to moderate concentrations of peptides or proteins (15-30 g L-1). As a result, the question of ACP-protein interactions in highly concentrated protein systems has clearly been overlooked, whereas it corresponds to actual industrial conditions such as drying or membrane filtration in the dairy industry for instance. In this study, the structure of an ACP phase is monitored in association with one model phosphorylated protein (casein) using solid-state nuclear magnetic resonance (ssNMR) under two conditions of high protein concentration (300 and 400 g L-1). At both concentrations and at 25 °C, it is found that the caseins maintain the mineral phase in an amorphous form with no detectable influence on its structure or size. Interestingly, and in both cases, a significant amount of the nonphosphorylated side chains interacts with ACP through hydrogen bonds. The number of these interacting side chains is found to be higher at the highest casein concentration. At 45 °C, which is a destabilizing temperature of ACP under protein-free conditions, the amorphous structure of the mineral phase is partially transformed at a casein concentration of 300 g L-1, while it remains almost intact at a casein concentration of 400 g L-1. Therefore, these results clearly indicate that increasing the concentration of proteins favors ACP-protein interactions and stabilizes the ACP clusters more efficiently.
authors.affiliation: Science et Technologie du Lait et de l'Œuf, INRA, UMR1253 , 65 rue de saint Brieuc, 35000 Rennes, France.|AGROCAMPUS OUEST, UMR1253 , 65 rue de saint Brieuc, 35000 Rennes, France.
authors.forename: Paulo
authors.identifier: 0000-0003-1004-3322
authors.initials: P
authors.lastname: De Sa Peixoto
authors.affiliation: Science et Technologie du Lait et de l'Œuf, INRA, UMR1253 , 65 rue de saint Brieuc, 35000 Rennes, France.|AGROCAMPUS OUEST, UMR1253 , 65 rue de saint Brieuc, 35000 Rennes, France.
authors.forename: Juliana V C
authors.initials: JV
authors.lastname: Silva
authors.affiliation: Sorbonne Universités , UPMC Univ Paris 6, CNRS, Collège de France, Laboratoire de Chimie de la Matière Condensée de Paris (LCMCP), Adresse, F-75005 Paris, France.
authors.forename: Guillaume
authors.initials: G
authors.lastname: Laurent
authors.affiliation: CAMPUS CNRS, Institut Charles Sadron , 23 rue du LOESS, BP 84047, 70034 Strasbourg, France.
authors.forename: Marc
authors.initials: M
authors.lastname: Schmutz
authors.affiliation: Team Translation and Folding, Université de Rennes 1, UMR CNRS 6290 IGDR , Campus de Beaulieu, 35000 Rennes, France.
authors.forename: Daniel
authors.initials: D
authors.lastname: Thomas
authors.affiliation: Science et Technologie du Lait et de l'Œuf, INRA, UMR1253 , 65 rue de saint Brieuc, 35000 Rennes, France.|AGROCAMPUS OUEST, UMR1253 , 65 rue de saint Brieuc, 35000 Rennes, France.
authors.forename: Antoine
authors.initials: A
authors.lastname: Bouchoux
authors.affiliation: Science et Technologie du Lait et de l'Œuf, INRA, UMR1253 , 65 rue de saint Brieuc, 35000 Rennes, France.|AGROCAMPUS OUEST, UMR1253 , 65 rue de saint Brieuc, 35000 Rennes, France.
authors.forename: Geneviève
authors.initials: G
authors.lastname: Gésan-Guiziou
mesh_terms: D002130:Calcium Phosphates / Q000737:chemistry*; D002364:Caseins / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D055550:Protein Stability; D013696:Temperature
chemical_list: D002130:Calcium Phosphates; D002364:Caseins; C519480:amorphous calcium phosphate