A cytoskeletal fraction of porcine tracheal smooth muscle (PTSM) was found to contain >90% of total cellular aldolase (fructose 1,6-bisphosphate aldolase, EC 4.1.2.13) activity. PTSM aldolase was purified by DEAE and inositol 1,4,5-trisphosphate (Ins(1,4,5)P) affinity chromatography and found to react with an antibody directed against human aldolase C, but not anti-aldolase A and B. The molecular mass of native aldolase was about 138 kDa (on Sephacryl S-300); SDS-denatured enzyme was 35 kDa (comigrated with rabbit skeletal muscle aldolase). Total cellular aldolase tetramer (aldolase) content was 34.5 pmol/100 nmol lipid P. Ins(1,4,5)P) binding activity coeluted with aldolase during Sephacryl 300, DEAE, and Ins(1,4,5)P affinity chromatography. Ins(1,4,5)P bound to purified aldolase (at 0 °C) in a dose-dependent manner over the range [Ins(1,4,5)P] 20 nM to 20 µM, with maximal binding of 1 mol of Ins(1,4,5)P/mol aldolase and a K of 12-14 µM. Fru(1,6)P and Fru(2,6)P displaced bound Ins(1,4,5)P) with a 50% inhibition at 30 and 170 µM, respectively. Ins(1,3,4)P (20 µM) and glyceraldehyde 3-phosphate (2 mM) were also potent inhibitors of Ins(1,4,5)P binding, but not inositol 4-phosphate or inositol 1,4-bisphosphate (20 µM each). Aldolase-bound Ins(1,4,5)P may play a role in phospholipase C-independent increases in free [Ins(1,4,5)P].

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