Formation of functional cross-species heterodimers of ornithine decarboxylase.
The two active sites in ornithine decarboxylase (ODC) are formed at the dimer interface with Lys-69 and Cys-360 contributing to each active site from opposite monomers [Tobias, K. E., & Kahana, C. (1993) Biochemistry 32, 5842-5847]. To gain insight into the organization of the substrate binding site and the nature of the dimer interface, analysis of ornithine decarboxylase from two parasitic protozoa, Trypanosoma brucei and Leishmania donovani, and from mouse was undertaken. Though T. brucei and mouse ornithine decarboxylase share only 60% sequence identity, the cross-species heterodimers form spontaneously, as measured by the restoration of enzyme activity upon mixing inactive K69A and C360A mutant enzymes. Thus, the amino acid composition of the dimer interface is apparently highly conserved between the T. brucei and mouse enzymes. Cross-species heterodimers were not formed between either T. brucei or mouse ODC and L. donovani ODC. Unlike the mouse and T. brucei ODC, the subunits of L. donovani ODC are not in rapid equilibrium, and incubation with a denaturant is required to induce reassociation. Kinetic analysis of the wild-type mouse and parasite ODCs revealed differences in the substrate binding sites between the three enzymes. The substrate binding properties of the restored active site in the T. brucei:mouse cross-species heterodimer mimic the characteristics of the wild-type enzyme from the species which contributes the subunit with a functional Lys-69.
Osterman
A
A
Department of Pharmacology, University of Texas Southwestern Medical Center, Dallas 75235.
Grishin
N V
NV
Kinch
L N
LN
Phillips
M A
MA
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D004926:Escherichia coli; D007700:Kinetics; D007893:Leishmania donovani / Q000201:enzymology; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D009838:Oligodeoxyribonucleotides; D009955:Ornithine Decarboxylase / Q000235:genetics / Q000378:metabolism*; D065108:Ornithine Decarboxylase Inhibitors; D011994:Recombinant Proteins; D013045:Species Specificity; D013379:Substrate Specificity; D014346:Trypanosoma brucei brucei / Q000201:enzymology
D009838:Oligodeoxyribonucleotides; D065108:Ornithine Decarboxylase Inhibitors; D011994:Recombinant Proteins; D009955:Ornithine Decarboxylase