Simultaneous binding of a polyamide dimer and an oligonucleotide in the minor and major grooves of DNA.

The effect of the polyamide ImPyPy-Dp (Im = N-methylimidazole-2-carboxamide, Py = N-methylpyrrole-2-carboxamide, and Dp = dimethylaminopropylamide), which binds as an antiparallel dimer in the Watson-Crick minor groove, on pyrimidine. purine.pyrimidine triple helix stability was investigated. A DNA restriction fragment was designed which contained two triple helix sites, one which overlapped a minor groove ligand site (proximal), and a control site 13 base pairs away (distal). Using quantitative DNase I footprint titration experiments the equilibrium association constant of oligonucleotide 5'-TTTTTm5CTTTm5CTTTm5CT-3' (1) to each site was measured in the absence and presence of the polyamide dimer. Our data indicate that triple helix formation is compatible with a polyamide dimer binding in the minor groove of DNA at an overlapping site. No cooperative effect of the polyamide dimer on the equilibrium association constant of oligonucleotide 1 was observed.

Cyclic polyamides for recognition in the minor groove of DNA.

Small molecules that specifically bind with high affinity to any designated DNA sequence in the human genome would be useful tools in molecular biology and potentially in human medicine. Simple rules have been developed to rationally alter the sequence specificity of minor groove-binding polyamides containing N-methylimidazole and N-methylpyrrole amino acids. Crescent-shaped polyamides bind as antiparallel dimers with each polyamide making specific contacts with each strand on the floor of the minor groove. Cyclic polyamides have now been synthesized that bind designated DNA sequences at subnanomolar concentrations.

Na+- and K+-dependent adenosine triphosphatase changes in multiple sclerosis plaques.

Na+- and K+-dependent adenosine triphosphatase [(Na+ + K+)-ATPase] was studied in microdissected samples from sections of histologically active and inactive plaques of multiple sclerosis. In active plaques the enzyme was found to be markedly reduced in activity, whereas in old, chronic plaques ATPase activities were higher than those of unaffected white matter. White matter from control patients and normal-appearing white matter from MS patients did not differ with regard to ATPase activities. The results suggest that reversible changes in the sodium pump mechanism may be involved in the pathophysiology of multiple sclerosis.

The ratio of 3-hydroxyacyl-CoA dehydrogenase to lipoamide dehydrogenase activity in individual muscle fibers: mitochondrial specialization for source of energy.

3-Hydroxyacyl-CoA dehydrogenase (HAD) has been widely used to assess the capacity for fatty acid oxidation by different muscle fiber types, with various Krebs cycle enzymes as a base for comparison. We have measured this enzyme in individual lyophilized fibers of the guinea pig soleus and the white and red portions of the vastus lateralis, and compared its activity in each fiber with that of lipoamide dehydrogenase (LAD), which as a part of the pyruvate dehydrogenase complex fulfills a function similar to HAD in forming acetyl-CoA, but from pyruvate and, thus, mainly from a carbohydrate source. The mean HAD/LAD ratio was 17.2 +/- 3 in the red vastus, 24.9 +/- 3 in the white vastus, and 43.7 +/- 10 in the soleus, all differences being highly significant. The two types of fast fibers were not distinguished from one another by the enzyme ratio within either the white or the red portion of the vastus lateralis. Data from all of the fast fibers taken together indicate a close correlation (0.93) between the two enzymes, whereas values from the soleus indicate a specialization of the mitochondria of the slow muscle fibers for the oxidation of fatty acids.

Fibrinolytic activity of plaques and white matter in multiple sclerosis.

Recent work has implicated plasminogen activator released from macrophages as a possible mediator of the demyelinating process in experimental allergic encephalomyelitis and multiple sclerosis (MS). We have studied the capacity of white matter and plaques from MS patients to break down fibrin clots, using a histochemical technique. Fibrinolytic activity was localized exclusively to areas around blood vessels and capillaries in both patients and controls. While there was marked variation between individuals, the unaffected white matter from MS patients was, on the average, not more active than that of controls, but plaques tended to show more numerous foci of lysis, often also more intense, than adjacent white matter; there was no correlation with disease activity or age of the plaques as determined by histological criteria. The localization and degree of fibrinolysis observed were not related to the presence of lymphocytic infiltrates, gliosis, or macrophages. However, the findings do not exclude an involvement of fibrinolytic enzymes (although originating from vascular endothelium rather than macrophages) in the genesis of the MS plaque, which commonly starts around a small vein.