Mannitol dehydrogenase (MtDH) is a key enzyme controlling the reductive synthesis of mannitol from fructose in the common mushroom Agaricus bisporus. A better understanding of the control of mannitol metabolism can be obtained by studying the structure of this enzyme. Here, the purification and crystallization of recombinant MtDH are reported. Crystals generally belonged to the space group C2, with unit-cell parameters a = 227, b = 125, c = 133 A, beta = 118 degrees, and diffracted to at least 1.8 A resolution, although a tantalum derivative belonged to the space group P2(1) and diffracted to the lower resolution of 2.9 A.