Purification and properties of a transketolase responsible for formaldehyde fixation in a methanol-utilizing yeast, candida boidinii (Kloeckera sp.) No. 2201.

Dihydroxyacetone synthase, present in methanol-grown Candida boidinii (Kloeckera sp.) No. 2201, catalyzes the transfer of the glycolaldehyde group from xylulose 5-phosphate to formaldehyde to form glyceraldehyde 3-phosphate and dihydroxyacetone. This enzyme was purified to electrophoretic homogeneity and found to be a new type of transketolase. The molecular weight of the enzyme was estimated to be 190,000 by gel filtration. The enzyme appeared to be composed of four identical subunits (Mr, 55,000). Thiamin pyrophosphate and Mg2+ were required for the activity. The optimum pH was found to be 7.0. With xylulose 5-phosphate as the ketol-donor, aliphatic aldehydes (C1-C7), glycolaldehyde and glyceraldehyde were better acceptors than ribose 5-phosphate. The kinetic data were consistent with a ping-pong bi-bi mechanism. The Km values obtained were as follows: xylulose 5-phosphate, 1.0 mM; formaldehyde, 0.43 mM; glyceraldehyde 3-phosphate, 0.42 mM; and dihydroxyacetone, 0.52 mM.