A multinuclear, high-resolution NMR study of bovine casein micelles and submicelles.
High-resolution, natural abundance 13C[1H] (100.5 MHz), 31P[1H] (161.8 MHz) and 1H (400.0 MHz) NMR spectroscopy was used to identify the calcium-binding sites of bovine casein and to ascertain the dynamic state of amino acid residues within the casein submicelles (in 125 mM KCl, pD = 7.4) and micelles (in 15 mM CaCl2/80 mM KCl, pD = 7.2). The presence of numerous, well-resolved peaks in the tentatively assigned 13C-NMR spectra of submicelles (90 A radius) and micelles (500 A radius) suggests considerable segmental motion of both side chain and backbone carbons. The partly resolved 31P-NMR spectra concur with this. Upon Ca2+ addition, the phosphoserine beta CH2 resonance (65.8 ppm vs DSS) shifts upfield by 0.2 ppm and is broadened almost beyond detection; a general upfield shift (up to 0.3 ppm) is also observed for the 31P-NMR peaks. The T1 values of the alpha CH envelope for submicelles and micelles are essentially identical corresponding to a correlation time of 8 ns for isotropic rotation of the caseins. Significant changes in the 31P T1 values accompany micelle formation. Data are consistent with a loose and mobile casein structure, with phosphoserines being the predominant calcium-binding sites.
Kakalis
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LT
U.S. Department of Agriculture, Eastern Regional Research Center, Philadelphia, PA 19118.
Kumosinski
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TF
Farrell
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HM
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002364:Caseins / Q000737:chemistry* / Q000378:metabolism; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D009682:Magnetic Resonance Spectroscopy; D008823:Micelles
D002364:Caseins; D008823:Micelles; D002118:Calcium