Lactoperoxidase-tubulin interactions.
Self-iodinated lactoperoxidase co-polymerizes with brain microtubules to constant specific activity and a stoichiometry of 0.2 to 0.3 lactoperoxidase molecule/tubulin heterodimer polymerized in the presence of 4 M glycerol. By contrast, iodinated tubulin loses its competence to polymerize. The lactoperoxidase-microtubule association is salt- and temperature-sensitive, shows considerable specificity, is saturable, and is reversible. Lactoperoxidase does not displace the microtubule-associated proteins from microtubules, does not promote polymerization, and binds to preformed microtubules. Self-iodinated lactoperoxidase also binds to tubulin oligomers at 0 degrees C and in the presence of CaCl2. The stoichiometry for this interaction is 0.6 to 0.8 molecules of enzyme/dimer. Lactoperoxidase forms a complex with soluble brain tubulin prepared by two cycles of polymerization and depolymerization or by phosphocellulose chromatography. The interaction was studied by sucrose gradient analysis, gel filtration, and spectral analysis based on the finding that tubulin binding to lactoperoxidase leads to a red shift in the Soret spectrum, yielding a difference spectrum with a minimum of 410 nm and maximum at 430 nm. This interaction involves one or more sulfhydryl groups of tubulin. Complex formation is relatively slow, is retarded by 0.6 M NaCl, and is accelerated by diiodotyrosine. By all three methods of analysis, the stoichiometry approaches a value of 2 lactoperoxidase molecules/tubulin dimer. There is a single class of binding sites in pig, beef, or rat tubulin with an apparent overall affinity constant of approximately 2.0 x 10(6) M-1. The molecular weight of the complex by sucrose gradient or gel filtration is approximately 140,000 i.e. half of the expected value for a 2:1 adduct. Since both alpha and beta subunits are present in the complex, we propose that the complex consists of a mixture of equal parts of presumably native alpha-tubulin-lactoperoxidase and beta-tubulin-lactoperoxidase.
D000818:Animals; D001921:Brain / Q000378:metabolism; D002417:Cattle; D007700:Kinetics; D007784:Lactoperoxidase / Q000378:metabolism; D046911:Macromolecular Substances; D008870:Microtubules / Q000378:metabolism; D010544:Peroxidases / Q000378:metabolism; D051381:Rats; D013053:Spectrophotometry; D013552:Swine; D014404:Tubulin / Q000378:metabolism
D046911:Macromolecular Substances; D014404:Tubulin; D007784:Lactoperoxidase; D010544:Peroxidases