Reversible dissociation/association of D-amino acid transaminase subunits: properties of isolated active dimers and inactive monomers.

The crystal structure of dimeric d-amino acid transaminase shows that the two Trp-139 sites are located in a hydrophobic pocket at the interface between the subunits and that the two indole side chains face one another and are within 10 A of coenzyme. This enzyme prefers an aromatic character at position 139, as previously demonstrated by the finding that Phe-139 but no other substitution tested provides the maximum degree of thermostability and catalytic efficiency. Here we show that an equilibrium between active dimers and inactive monomers can be demonstrated with the W139F mutant enzyme, whereas with the wild-type enzyme the subunit interface is so tight that a study of this equilibrium is precluded. We show how the processes of dimerization of monomers and dissociation of dimers to monomers are controlled. Lower pH (5.0) favors monomer formation from dimers. Gel filtration and activity analysis show that at higher pH (7.0) the monomers combine to form active dimers with a Kd of 0.17 μM. This assembly...