Extended Data Fig. 2: PLP regulates SHMT2 dimer-tetramer transition.

a, PLP association with the indicated forms of SHMT2ΔN. SHMT2–PLP internal aldimine linkage was monitored by measuring absorbance at 435 nm after rapid mixing of SHMT2ΔN (5 μM) and PLP (100 μM). Data are mean ± s.e.m. of four independent reactions. b, Enzyme activity of the indicated forms of SHMT2ΔN (0.5 μM) against the synthetic substrate l-threo-phenylserine in the presence of 50 μM PLP. The reaction was monitored by measuring benzyldehyde absorbance at 279 nm. Data are mean ± s.e.m. of three independent experiments carried out in duplicate. c, Differential scanning fluorimetry analysis of different forms of SHMT2ΔN in the presence and absence of PLP. SHMT2ΔN wild-type and mutant proteins (5 μM) were incubated with the indicated concentrations of PLP or 100 μM 2-hydroxy-6-methylpyridine-3-carboxylic acid (HMCA) for 10 min at 20 °C before differential scanning fluorimetry analysis. d, Changes in melting temperatures (ΔT_m) for each of the conditions shown in c. ΔT_m was calculated by subtracting the T_m of SHMT2 with buffer without ligands from the T_m of SHMT2 with added ligands. Data in c, d represent a single experiment carried out in duplicate. e, Summary of SHMT2ΔN(A285T) dimer–tetramer equilibrium in response to PLP binding.