FIG. 4.

PAPS biosynthesis in nodPQ, cysH, and nodS mutants. Formic acid-extractable material was fractionated by thin-layer chromatography, and the incorporation of ³⁵SO₄ was visualized by autoradiography as described in Materials and Methods. (A) In vitro-synthesized ³⁵S-labeled PAPS was added to 40 μl of an unlabeled M. loti formic acid extract and spotted onto a TLC plate (lane 1). Wild-type M. loti (lane 2), wild-type M. loti harboring either empty vector (lane 3) or pGTO101 (plasmid carrying nodPQ) (lane 4), the nodPQ mutant (lane 5), the nodPQ mutant harboring either empty vector (lane 6) or pGTO101 (lane 7), the cysH mutant (lane 8), and the nodS mutant (lane 9) were cultured in minimal medium in the presence of Na₂³⁵SO₄ for 48 h. (B) E. coli mutants were cultured in minimal medium in the presence of Na₂³⁵SO₄ for 12 h, and formic acid extracts were examined as for panel A. Lane 1, PAPS; lane 2, wild type (MG1655); lane 3, cysN (DM63); lane 4, cysH (JM96); lane 5, cysI (CAG12182); lane 6, cysJ (AT2427); lane 7, cysK (RL165).