MPTP, MPP+ and mitochondrial function

1-Methyl-4-phenylpyridinium (MPP+), the putative toxic metabolite of the neurotoxin, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), inhibited NAD(H)-linked mitochondrial oxidation at the level of Complex I of the electron transport system. MPTP and MPP+ inhibited aerobic glycolysis in mouse striatal slices, as measured by increased lactate production; MPTP-induced effects were prevented by inhibition of monoamine oxidase B activity. Several neurotoxic analogs of MPTP also form pyridinium metabolites via MAO; these MPP+ analogs were all inhibitors of NAD(H)-linked oxidation by isolated mitochondria. 2'-Methyl-MPTP, a more potent neurotoxin in mice than MPTP, was also more potent than MPTP in inducing lactate accumulation in mouse brain striatal slices. Overall, the studies support the hypothesis that compromise of mitochondrial oxidative capacity is an important factor in the mechanisms underlying the toxicity of MPTP and similar compounds.     

The structure of toxic protein, the mistletoe lectin, at different pH: the study using intrinsic fluorescence method

The effects of pH on the conformation of mistletoe lectin I and its isolated A- and B-subunits has been investigated by using the methods of intrinsic fluorescence. By the denaturating action of guanidine hydrochloride and the influence of the quenchers (I-, Cs+, acrylamide) the structural stability of the native protein and its isolated subunits was estimated. Treatment of the protein with the denaturant and quenchers revealed its different structure at pH 7.0 and 4.0. At pH 4.0 tryptophan residues become more accessible to quenchers, positive charge of the surrounding area increases and the protein becomes more stable to the action of denaturant. The structure of the isolated A- and B-chains of mistletoe lectin I differs considerably from that of the whole protein: a) its stability to the action of guanidine hydrochloride is lower; b) it depends on the ionic strength of the solvent; c) it is characterized by increased accessibility of tryptophan residues to quenchers (for B-chain). Differences between the conformations of the isolated chains at pH 7.0 and 4.0 are marked more strongly; moreover, at pH 4.5 the B-chain undergoes structural transition. The possible relationship between structural peculiarities of mistletoe lectin I and the mechanism of its transmembrane transfer is discussed.     

A variable region gene subfamily encoding T cell receptor beta-chains is selectively conserved among mammals

Studies of murine T cell receptor genes indicate that the VT beta repertoire, in contrast to the large VT alpha, VH, and VK repertoires, is limited to approximately 21 genes. Large differences between the various VT beta sequences allow classification into distinct subfamilies consisting of one or few members. The VT beta sequence of a gene, RTB92, expressed in a rabbit T cell line is most closely related to a VT beta gene expressed in the human cell line Molt-4. Genomic blots with RTB92 VT beta region used as probe indicated conservation of related VT beta gene subfamilies in man and pig, but no related sequences were seen in rat, mouse, or hamster. The relationship among these VT beta genes mirrors similarities and differences in MHC genes of the compared species and suggests coordinate evolution of these functionally related gene families. The constant region of RTB92 was shown to be rabbit CT beta 2 by reference to genomic clones. Comparisons with constant region sequences of human and murine CT beta 1 and CT beta 2 chains reveal no similarities in amino acid or nucleic acid sequence in the coding region characteristic of the respective isotypes. Intraspecies homologies between CT beta 1 and CT beta 2 sequences were much greater than those between CT beta 1 or CT beta 2 from other species. By contrast, comparisons of JT beta and 3' untranslated region sequences showed significant interspecies conservation of sequences in the beta 1 and in the beta 2 regions.     

Covalent alteration of the prosthetic heme of human hemoglobin by BrCCl3. Cross-linking of heme to cysteine residue 93.

Recent studies have shown that a protein-bound heme adduct formed from the reaction of BrCCl3 with myoglobin was due to bonding of the proximal histidine residue through the ring I vinyl of a heme-CCl2 moiety. The present study reveals that BrCCl3 also reacts with the heme of reduced human hemoglobin to form two protein-bound heme adducts. Edman degradation and mass spectrometry provided evidence that these protein-bound heme adducts were addition products in which heme-CCL2 or heme-CCl3 were bound to cysteine residue 93 of the beta-chain of hemoglobin. It appeared that the cysteine residue was bonded regiospecifically to the ring I vinyl group of the altered heme moiety, because the nonprotein-bound products of the reaction included the beta-carboxyvinyl and alpha-hydroxy-beta-trichloromethylethyl derivatives of the ring I vinyl moiety of heme. The absorption spectra of the protein-bound adducts in both the oxidized and reduced states were highly similar to those described for hemichromes, which are thought to be involved in the formation of Heinz bodies and subsequent red cell lysis.     

Serologic and structural characterization of immunoglobulin chains secreted by rabbit-mouse hybridomas

Serologic and primary structural analyses of Ig chains secreted by several rabbit-mouse hybridomas have shown that these hybrid cells produce heavy (H) or light (L) chains identical to those isolated from rabbit sera. Two of the cell lines (7D2, 7D6) secreted rabbit H chains with a m.w. of 55,000 each of which expressed a full complement of variable and constant region allotypes (a3, d11, e15). These apparently normal rabbit H chains were secreted in a complex with a m.w. about 130,000, and serologic studies indicated that this complex contained a covalently linked mouse kappa L chain. Two other cell lines (4C1, 12F2) produced allotype b4 L chains with m.w. of 23,000 and 25,000, and a third (1D4P5) produced an allotype b5 L chain with a m.w. of 23,000. Serologic analyses indicated that the allotypes on these chains are equivalent to those expressed by normal rabbit Ig molecules. Partial amino acid sequence data obtained for the L chain products showed them to be typical of rabbit L chains, and to be significantly different from mouse L chains.     

Prevention of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced dopaminergic toxicity in mice by MDL 72145, a selective inhibitor of MAO-B

Pretreatment of mice with the potent and selective monoamine oxidase B (MAO-B) inhibitor MDL 72145 ((E)-2-(3',4'-dimethoxyphenyl)-3-fluoroallylamine) protected against the dopaminergic neurotoxicity of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Mice treated with MDL 72145 prior to MPTP did not exhibit the decrement in the neostriatal content of dopamine and its metabolites normally seen after MPTP administration. This observation adds further support to the concept that the oxidation of MPTP by MAO-B to its corresponding pyridinium analog, 1-methyl-4-phenylpyridinium (MPP+), is an important feature of the neurotoxic process.