ACTIVE MONOMERIC AND DIMERIC FORMS OF PSEUDOMONAS-PUTIDA GLYOXALASE-I- EVIDENCE FOR 3D DOMAIN SWAPPING

3D domain swapping of proteins involves the interconversion of a monom er containing a single domain-domain interface and a 2-fold symmetrica l dimer containing two equivalent intermolecular interfaces. Human gly oxalase I has the structure of a domain-swapped dimer [Cameron, A. D., Olin, B., Ridderstrom, M., Mannervik, B., and Jones, T. A. (1997) EMB O J. 16, 3386-3395] but Pseudomonas putida glyoxalase I has been repor ted to be monomeric [Rhee, H.-L, Murata, K., and Kimura, A. (1986) Bio chem. Biophys. Res. Commun. 141, 993-999]. We show here that recombina nt P. putida glyoxalase I is an active dimer (k(cat) similar to 500 +/ - 100 s(-1); K-M similar to 0.4 +/- 0.2 mM) with two zinc ions per dim er. The zinc is required for structure and function. However, treatmen t of the dimer with glutathione yields an active monomer (k(cat) simil ar to 115 +/- 40 s(-1); K-M similar to 1.4 +/- 0.4 mM) containing a si ngle zinc ion. The monomer is metastable and slowly reverts to the act ive dimer in the absence of glutathione. Thus, glyoxalase I appears to be a novel example of a single protein able to exist in two alternati ve domain-swapped forms. It is unique among domain-swapped proteins in that the active site and an essential metal binding site are apparent ly disassembled and reassembled by the process of domain swapping. Fur thermore, it is the only example to date in which 3D domain swapping c an be regulated by a small organic ligand.