Purification and Characterization of a Human Brain Galectin-1 Ligand

Abstract: Our previous studies have characterized an endogenous lectin from human brain identified as galectin-1. A soluble ligand of galectin-1 was purified from human brain by affinity chromatography and preparative electrophoresis. The purified ligand (termed HBGp82, for human brain galectin-1-binding polypeptide of 82,000 daltons) has an apparent molecular mass of 82 kDa and is glycosylated by N -linked biantennary complex structures. HBGp82 was partially characterized by microsequencing of peptide fragments. Similar peptides were found in a heat shock of protein of 90,000 daltons, hsp90. However, comparison of apparent molecular weights and matrix-assisted laser desorption mass spectrometry clearly showed that HBGp82 differs to some degree from hsp90.     

Model of synchronized population bursts in electrically coupled interneurons containing active dendritic conductances

We constructed a computer model of 128 interneurons, each with multiple dendritic branches and an axonal segment. The model neurons were interconnected by gap junctions between dendritic compartments, as are known to occur in rat and guinea-pig hilar interneurons. The model contained no excitatory synapses. In the presence of low-frequency spontaneous action potentials, the model generated synchronized population bursts, when gap junction resistance was 50 M and there were at least two gap junctions per neuron on average. Population bursts occurred only when the dendrites of model neurons were electrically excitable. Consistent with experiment, somatic hyperpolarization during the population burst uncovered partial spikes. In the model, partial spikes originated in electrically active dendrites driven by coupled dendrites. This model may account for population bursts in hilar interneurons that occur in 4-aminopyridine (4AP) together with blockers of GABA_A and excitatory amino acid (EAA) receptors.     

Multiple paternity in the red-eyed treefrog Agalychnis callidryas (Cope)

DNA fingerprinting of tadpoles from two different ‘one female-two male’ matings of the red-eyed treefrog, Agalychnis callidryas, revealed multiple paternity of offspring. Offspring were assigned paternity based on bands shared with the putative father, but not shared between putative fathers or with the mother. Paternity was split 44/56% and 36/64% in two matings. These results do not support a hypothesis of sperm priority in access to unfertilized eggs by primary males. Multiple paternity may be commonplace in species of anurans with matings by multiple males.     

Steady-state kinetics of the overall oxidative phosphorylation reaction in heart mitochondria. Determination of the coupling relationships between the respiratory reactions and miscellaneous observations concerning rate-limiting steps

The linear sequence of steps involved in the oxidation of extramitochondrial succinate by O₂ in bovine heart mitochondria was examined by a steady-state kinetic method to determine whether or not freely diffusible intermediates occur between the various inhibitor-sensitive steps. The kinetic method is based on the facts (1) that if two inhibitor-sensitive steps within a sequence are linked by a freely diffusible intermediate, inhibition of one will make the other less rate limiting in the overall reaction and thus will increase the amount of inhibitor of the other step required for half-maximal inhibition of the overall reaction, and (2) that if the two steps are not linked in this manner, inhibition of one will make the other more rate limiting and thus will decrease the amount of inhibitor of the other required for half-maximal inhibition. These two types of coupling relationships between steps were designated as sequential and fixed, respectively. The results indicate the existence of freely diffusible intermediates (sequential coupling relationships) between the succinate transport and succinate dehydrogenase reactions, between the succinate dehydrogenase and cytochromebc ₁ reactions, and between the cytochromesbc ₁ andaa ₃ reactions. Uncoupling respiration from phosphorylation results in the coupling relationship between thebc ₁ andaa ₃ reactions becoming partially fixed. This change is accompanied by marked decreases in the degrees to which thebc ₁ andaa ₃ reactions limit the overall reaction and appears to account for the large uncoupler-induced releases of inhibition at the levels of thebc ₁ andaa ₃ reactions observed previously by others. It is suggested that cytochromec is the freely diffusible intermediate between thebc ₁ andaa ₃ reactions and that the uncoupler-induced changes occur as a result of formation of functional and highly efficient supercomplexes between cytochromec and the cytochromesbc ₁ andaa ₃ complexes.     

Purification and characterization of two essential cytochromes of the thiosulphate-oxidizing multi-enzyme system from Thiobacillus A2 (Thiobacillus versutus)

Four c-type cytochromes were purified by several procedures including chromatography on DEAE-Sepharose CL-6B, Phenyl-Sepharose CL-4B and Sephadex G-75, G-100 and G-200 and chromatofocusing. Cytochrome c-551 had a pI value of 5.2 and an M_r of 260 000 consisting of six non-covalently bound polypeptides each with an M_r of 43 000, and contained four to five haems. Cytochrome c-552.5 had a pI value of 4.8 and an M_r of 56 000 consisting of two polypeptides with the same M_r 29 000, and contained two haems. Cytochromes c-551 and c-552.5 were reduced by ascorbate to about 70 and 60% of the fully dithionite-reduced values, respectively, and both were essential components in the thiosulphate-oxidizing multi-enzyme system (other components of the system were ‘enzyme A’, ‘enzyme B’ and sulphite: cytochrome c oxidoreductase). These two cytochromes functioned as electron carriers and effectors in the oxidation of thiosulphate. Some evidence suggested that cytochrome c-551 might be a specialized electron transfer component for sulphonate-sulphur oxidation. Both cytochromes could be reduced by thiosulphate in the presence of enzymes A and B. Cytochrome c-550 (basic) and cytochrome c-550 (acidic) were small proteins with M_r 15 000 and 14 000 and pI values of over 8 and 5, respectively. Their physiological role is uncertain.     

Incorporation of [14C]cinnamate into hydrolase-resistant components of the primary cell wall of spinach

[¹⁴C]Cinnamate was taken up very rapidly by cultured spinach cells and completely incorporated into low-MW conjugates within 20 min. The ¹⁴C-labelled products were similar whether the [¹⁴C]cinnamate was supplied continuously over a period of hours via a peristaltic pump or instantaneously. Radioactivity was slowly recruited from the low-MW pool into aromatic components of the cell-wall fraction. Saponification of the radioactive wall fraction yielded, in addition to radioactive ferulate and p-coumarate, large amounts of ethyl acetate-soluble radioactive material with the properties of oxidatively coupled phenols. The coupled material was associated with the most highly ‘Driselase’-resistant fractions of the cell wall. In contrast, ‘Driselase’ released most of the wall's ferulate and p-coumarate on disaccharide fragments. It is suggested that the oxidatively coupled phenols are formed from simpler phenols by peroxidase and that they cross-link the polysaccharides to which they are attached, making these polysaccharides relatively ‘Driselase’-resistant.     

Vascular permeability to proteins and peptides in the mouse pineal gland

Summary The permeability of fenestrated capillaries in the mouse pineal gland to proteins and peptides was demonstrated by means of ultrastructural tracers. Horseradish peroxidase (HRP) and microperoxidase (MP) were injected intravenously and allowed to circulate for approximately 30 s, 1 min, 5 min, 1 or 2h. The tissue was then fixed by vascular perfusion or by immersion with aldehydes. In all experiments a pronounced extravasation of HRP and MP occurred. Transendothelial vesicular transport seemed to have occurred across the fenestrated capillaries. The most pronounced tracer labeling of vesicles was found after 1 min of MP- or HRP-circulation. The vesicles were uncoated and more than 70 % of the HRP-and MP-containing vesicles exhibited diameters between 50 and 110 nm. Furthermore, three other transcapillary pathways taken by the tracers are suggested: 1) via intercellular junctions, 2) through fenestrae and 3) via channels formed by fusion of vesicles with the luminal and abluminal cell membranes. Based on these results, it is assumed that the capillaries in the mouse pineal gland are also permeable to peptides synthesized and secreted by the pineal gland.Part of this study was presented at the EMCELL-76 meeting, Copenhagen, 1976     

Spatial distribution of seston in an artificially mixed system

Seston size distribution was determined with a Coulter Counter to investigate seston interaction and its effects on phytoplankton production. Spatial distribution patterns of the non-productive component of the seston were as complex as those commonly associated with algae. Seston might interact influencing production by changes in optical depth, abrasive actions and nutrient adsorption.