FIGURE 2.
A, products of the SPT reaction using serine, alanine, or glycine as substrates. The conjugation of palmitoyl-CoA with alanine and glycine leads to the formation of the two DSBs: m18:0 and m17:0. B, chemical structure of the DSBs. An abbreviated nomenclature for sphingoid bases is used in this work. The numbers of hydroxyls are designated by m (for mono-) and d (for di-) followed by the number of carbons. The second number indicates the double bonds. For example, d18:0 stands for sphinganine, and d18:1 stands for sphingosine. All shown metabolites were also found in the N-acetylated form. C, accumulation of m18:0 and m17:0 in HEKC133W cells after supplementing the culture medium with alanine or glycine. HEKC133W cells were cultured using either standard medium (−) or medium that was supplemented with 10 mm alanine (+ala) or 10 mm glycine (+gly). De novo synthesis was blocked with FB1 for 24 h, and the accumulated lipids were analyzed by LC-MS. D, accumulation of DSB in HEK cells expressing mutant forms of SPT. HEKempty, HEKL1, HEKC133W, and HEKC133Y cells were treated with FB1 for 24 h, and the extracted lipids were quantified by LC-MS. Error bars in C and D indicate S.E. p values ≤ 0.01 were labeled with **.