FIGURE 8.

Activity and acyl-CoA selectivity of monomeric and triple-fusion SPTs in yeast and CHO-Ly-B cells. A, the hLCB1 and hLCB2a genes were expressed in lcb1Δlcb2Δtsc3Δ yeast cells along with wild-type ssSPTa or ssSPTb as described under “Experimental Procedures.” Alternatively, single-chain hLCB2a-ssSPTa-hLCB1 or hLCB2a-ssSPTb-hLCB1 fusions were expressed in yeast cells to ensure stoichiometric expression of the three subunits. Microsomal SPT activities were measured as described under “Experimental Procedures” using either C16- or C18-CoA as substrate. B, monomeric subunits (left) or the triple fusions (right) containing the indicated ssSPTs were expressed in yeast. LCBs were prepared and analyzed by HPLC as described under “Experimental Procedures.” C, monomeric subunits or triple fusions containing the indicated ssSPTs were expressed in CHO-Ly-B cells, and microsomal SPT activities were determined as described under “Experimental Procedures.”