Active Monomeric and Dimeric Forms of Pseudomonas putida Glyoxalase I:  Evidence for 3D Domain Swapping

• Published in: Biochemistry (Easton) Vol. 37; no. 29; pp. 10345 - 10353
• Main Authors: Saint-Jean, André P, Phillips, Kristina R, Creighton, Donald J, Stone, Martin J
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 21.07.1998
• Subjects: Apoenzymes - chemistry; Apoenzymes - isolation & purification; Binding Sites; Dimerization; Enzyme Stability - drug effects; Glutathione - pharmacology; Humans; Lactoylglutathione Lyase - chemistry; Lactoylglutathione Lyase - genetics; Lactoylglutathione Lyase - metabolism; Models, Molecular; Protein Structure, Secondary; Protein Structure, Tertiary; Pseudomonas putida; Pseudomonas putida - enzymology; Recombinant Proteins - biosynthesis; Recombinant Proteins - chemistry; Zinc - chemistry
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/bi980868q

3D domain swapping of proteins involves the interconversion of a monomer containing a single domain−domain interface and a 2-fold symmetrical dimer containing two equivalent intermolecular interfaces. Human glyoxalase I has the structure of a domain-swapped dimer [Cameron, A. D., Olin, B., Ridderström, M., Mannervik, B., and Jones, T. A. (1997) EMBO J. 16, 3386−3395] but Pseudomonas putida glyoxalase I has been reported to be monomeric [Rhee, H.-I., Murata, K., and Kimura, A. (1986) Biochem. Biophys. Res. Commun. 141, 993−999]. We show here that recombinant P. putida glyoxalase I is an active dimer (k cat ∼500 ± 100 s-1; K M ∼0.4 ± 0.2 mM) with two zinc ions per dimer. The zinc is required for structure and function. However, treatment of the dimer with glutathione yields an active monomer (k cat ∼115 ± 40 s-1; K M ∼1.4 ± 0.4 mM) containing a single zinc ion. The monomer is metastable and slowly reverts to the active dimer in the absence of glutathione. Thus, glyoxalase I appears to be a novel example of a single protein able to exist in two alternative domain-swapped forms. It is unique among domain-swapped proteins in that the active site and an essential metal binding site are apparently disassembled and reassembled by the process of domain swapping. Furthermore, it is the only example to date in which 3D domain swapping can be regulated by a small organic ligand.