L phenylalanine 13c9

U-¹³C Tyr-(D)Ala-Phe (YAF) tripeptide was synthesized using labeled amino acids purchased from Sigma-Aldrich (D-Alanine-¹³C₃-99%) and Cambridge Isotope Laboratories, Inc. (L-Tyrosine-¹³C₉-99%, ¹⁵N-99% and L-Phenylalanine-¹³C₉-99%, ¹⁵N-99%). Its synthesis was carried out in solution according to procedure reported in our previous paper.²³ U-¹³C,¹⁵N-GB1 was bought from Giotto Biototech company as “ready-to-use” sample packed in 1.3 mm rotor. Dynein Light Chain 8 (LC8) protein was prepared according to the protocol reported by us previously.^{24 (link)} 3 mg of U-¹³C,¹⁵N-LC8 prepared by controlled precipitation from polyethylene glycol were packed into a 1.3 mm rotor and used for subsequent MAS NMR experiments.

Internal standards included the following stable-isotope labeled compounds: [24-¹³C]-cholic acid, l-Arginine-¹⁵N4, Uracil-¹⁵N2, l-Tyrosine-¹³C9,¹⁵N, l-Phenylalanine-¹³C9, ¹⁵N, and Acetyl-d3-Carnitine purchased from Cambridge Isotope laboratories; linoleic acid-d11 and eicosapentaenoic acid-d5 purchased from Cayman Chemicals; and [trimethyl-13C3]-caffeine purchased from Milli-poreSigma. Acetonitrile and formic acid (Optima, Fisher Scientific) and isopropyl alcohol (Lichrosolv, MillioporeSigma) used for analysis were LC–MS grade. Water was prepared with an Ultrapure MilliQ system (18 MΩ, Millipore, USA). Purified human hemoglobin (MP Biomedicals), Human Serum Albumin (MilliporeSigma), and Sysmex Reagent Sulfolyser (Henry Schein, Melville, NY) were purchased for the Hb assays.