Structural comparison of the 68 kDa laminin-binding protein and 5'-nucleotidase from chicken muscular sources: evidence against a gross structural similarity of both proteins

The 68 kDa laminin-binding protein purified from chicken skeletal muscle and the ectoenzyme 5'-nucleotidase from chicken gizzard are both able to interact with laminin. They were both shown to possess a nearly identical amino acid composition. The 79 kDa glycosylated form of 5'-nucleotidase can be transformed into an enzymatically active form by treatment with endoglycosidase F (Endo F). Deglycosylated (Endo F-treated) 5'-nucleotidase exhibits an apparent molecular mass of 68 kDa. Using immunological and finger-printing techniques, both proteins were analysed to determine their structural relatedness. The results obtained indicate that both proteins are not identical but may posses a few common peptides of yet unknown sequence and length.     

Angle of the retinal of bacteriorhodopsin in blue membrane

The electric dichroism of purple and cation-depleted (blue) membrane was measured in a.c. electric fields at saturation. A decrease of 5.5° in the direction of the chromophore transition moment with respect to the membrane normal was found upon removal of cations from purple membrane.     

Temperature jump study of charge translocation during the bacteriorhodopsin photocycle

Temperature jump experiments were carried out on purple membranes oriented and fixed in polyacrylamide gel. With green background illumination a relaxation of the photocurrent after an infrared laser pulse could be observed. To simulate the temperature jump signals different models of the bacteriorhodopsin photocycle were tested. The parameters of these models were obtained by measuring absorbance changes and photocurrent after excitation with a 575-nm laser flash.

A model with a temperature-dependent branching before the M state turned out to be satisfying. Other models, especially those with a late branching or without branching, could not reproduce the temperature jump measurements.

     

Incorporation of ionic channels from yeast plasma membranes into black lipid membranes.

Recently, patch-clamping of yeast protoplasts has revealed the presence of plasma membrane K+ channels (Gustin, M. C., B. Martinac, Y. Saimi, M. R. Culberston, and C. Kung. 1986. Science (Wash. DC). 233:1195-1197). In this work we show that fusion of purified plasma membranes into planar bilayers allows the study of the yeast channels. The main cationic conductances detected were of 64 and 116 pS, however, larger and smaller conductances have been observed. The two main conductances were sensitive to the K+ channels blockers tetraethylammonium (TEA+) and Ba2+. Bionic experiments indicated that both conductances were K+ selective.     

Photoreactions of bacteriorhodopsin at acid pH.

It has been known that bacteriorhodopsin, the retinal protein in purple membrane which functions as a light-driven proton pump, undergoes reversible spectroscopic changes at acid pH. The absorption spectra of various bacteriorhodopsin species were estimated from measured spectra of the mixtures that form at low pH, in the presence of sulfate and chloride. The dependency of these on pH and the concentration of Cl- fit a model in which progressive protonation of purple membrane produces "blue membrane", which will bind, with increasing affinity as the pH is lowered, chloride ions to produce "acid purple membrane." Transient spectroscopy with a multichannel analyzer identified the intermediates of the photocycles of these altered pigments, and described their kinetics. Blue membrane produced red-shifted KL-like and L-like products, but no other photointermediates, consistent with earlier suggestions. Unlike others, however, we found that acid purple membrane exhibited a very different photocycle: its first detected intermediate was not like KL in that it was much more red-shifted, and the only other intermediate detectable resembled the O species of the bacteriorhodopsin photocycle. An M-like intermediate, with a deprotonated Schiff base, was not found in either of these photocycles. There are remarkable similarities between the photoreactions of the acid forms of bacteriorhodopsin and the chloride transport system halorhodopsin, where the Schiff base deprotonation seems to be prevented by lack of suitable aspartate residues, rather than by low pH.     

Sliding controller design for a nonlinear fermentation system

The design of a sliding controller for a continuous fermentation process is presented. The results obtained by simulation have proved the control scheme to be very robust. Regulation of substrate concentration at its optimal value has been achieved even though process parameters change their nominal values. Chattering effects are decreased by introducing a minor modification of the control variable around the sliding surface.     

Expression of nonspecific cross-reacting antigen species in myeloid leukemic patients and healthy subjects

The reactivity of two monoclonal antibodies recognizing NCA-95 and NCA-55 (MAb 47 and MAb 192, respectively) with a polyclonal anti-NCA serum in myeloid leukemic cells isolated by density gradient centrifugation was compared using an immunofluorescence test (IF). It was observed that the blood myeloid cells in 78.8% of the patients with different types of myelocytic leukemias and all granulocytes of 15 normal donors showed similar expression of the NCA species studied. The leukocytes of the remaining patients did not synthesize the NCA-95 species regardless of the maturation stage of the cells studied. In two patients, synthesis of this NCA form was limited to the fractions containing myelocytes and metamyelocytes. We have found that all anti-NCA antibodies studied recognized different antigenic epitopes in a myeloid cell series. A relationship between the patient's survival and the proportion of NCA-containing cells was also observed.     

Chronic immune thrombocytopenic purpura--immunological analyses of a patient pre- and post-HIV seroconversion

A seven year follow-up of immune parameters is reported for a patient with chronic immune thrombocytopenic purpura (ITP) pre and post human immunodeficiency virus (HIV) seroconversion. Therapies such as intravenous IgG, prednisone, vincristine, or Ciclosporin A had no clear-cut beneficial effect on platelet counts. A long-term normalization of platelet counts was achieved by splenectomy. At splenectomy the patient was seropositive for HIV, most likely transmitted by blood products received half a year prior to laparatomy. Mean plasma levels of the second component of complement, C2, were half of the normal values prior to and within the lowest normal range post HIV seroconversion. Nevertheless, the T cell-dependent B cell response to HIV, which is dependent on the activation of C3 via the classical pathway of complement, was normal: Western blot analysis of total IgG and of IgG subclass responses to individual HIV antigens proved to be unimpaired.     

The use of functional analysis of the ribosome as a tool to determine archaebacterial phylogeny

Forty different antibiotics with diverse kingdom and functional specificities were used to measure the functional characteristics of the archaebacterial translation apparatus. The resulting inhibitory curves, which are characteristic of the cell-free system analyzed, were transformed into quantitative values that were used to cluster the different archaebacteria analyzed. This cluster resembles the phylogenetic tree generated by 16S rRNA sequence comparisons. These results strongly suggest that functional analysis of an appropriate evolutionary clock, such as the ribosome, is of intrinsic phylogenetic value. More importantly, they indicate that the study of the nexus between genotypic and phenotypic (functional) information may shed considerable light on the evolution of the protein synthetic machinery.     

Inhibition of carbonic anhydrases in type I muscle fibers influences contractility

We tested the effects of inhibiting the carbonic anhydrase activity of rat soleus and extensor digitorum longus muscles on the isometric contractile properties and the resistance to fatigue. SOL and EDL muscles from female rats were incubated in vitro in the presence of methazolamide, a specific inhibitor of carbonic anhydrase, before determining their contractile properties. Methazolamide had no effects on the contractile properties of the soleus muscle (10(-5) or 10(-3) M) and extensor digitorum longus (10(-3) M), except for the half-relaxation time of the soleus muscle which increased significantly. Values for half-relaxation time were significantly increased with both concentrations of the inhibitor. Muscles were then submitted to a fatigue protocol lasting 30 min. During the fatigue test, no significant difference was observed between control and 10(-5) M methazolamide soleus muscles. In presence of 10(-3) M methazolamide however, the soleus muscle showed a significantly increased resistance to fatigue compared with control preparations. No significant effect was observed with the extensor digitorum longus muscle exposed to 10(-3) M methazolamide. Results are discussed in terms of the presence of two different isoforms of carbonic anhydrase that may be associated with calcium uptake and energy metabolic processes, respectively.