Design, synthesis, and characterization of a photoactivatable flavocytochrome molecular maquette

We report the construction of a synthetic flavo-heme protein that incorporates two major physiological activities of flavoproteins: light activation of flavin analogous to DNA photolyase and rapid intramolecular electron transfer between the flavin and heme cofactors as in several oxidoreductases. The functional tetra-alpha-helix protein comprises two 62-aa helix-loop-helix subunits. Each subunit contains a single cysteine to which flavin (7-acetyl-10-methylisoalloxazine) is covalently attached and two histidines appropriately positioned for bis-his coordination of heme cofactors. Both flavins and hemes are situated within the hydrophobic core of the protein. Intramolecular electron transfer from flavosemiquinone generated by photoreduction from a sacrificial electron donor in solution was examined between protoporphyrin IX and 1-methyl-2-oxomesoheme XIII. Laser pulse-activated electron transfer from flavin to meso heme occurs on a 100-ns time scale, with a favorable free energy of approximately -100 meV. Electron transfer from flavin to the lower potential protoporphyrin IX, with an unfavorable free energy, can be induced after a lag phase under continuous light illumination. Thus, the supporting peptide matrix provides an excellent framework for the positioning of closely juxtaposed redox groups capable of facilitating intramolecular electron transfer and begins to clarify in a simplified and malleable system the natural engineering of flavoproteins.     

Long-term progression in chronic manganism: ten years of follow-up

We studied the long-term clinical course of five patients with chronic manganese intoxication. The mean scores of the King's College Hospital Rating Scale for Parkinson's disease increased from 15.0 +/- 4.2 in 1987 to 28.3 +/- 6.70 in 1991 and then to 38.1 +/- 12.9 in 1995. The deterioration was most prominent in gait, rigidity, speed of foot tapping, and writing. Tissue concentrations of manganese in blood, urine, scalp hair, and pubic hair returned to normal. Follow-up MRIs did not show paramagnetic high-signal intensity on T1-weighted images. The data indicate that clinical progression in patients with manganese parkinsonism continues even 10 years after cessation of exposure.     

Transient characteristics of small salient-pole alternators

The use of small alternators, other than for standby purposes, is increasing and this has focused attention on the reliability and utility of the parameters normally quoted for such machines. Test procedures are described and methods used for parameter estimation discussed, with particular emphasis on the difficulties presented by small brushless machines     

Activation of auditory cortex during silent lipreading

Watching a speaker's lips during face-to-face conversation (lipreading) markedly improves speech perception, particularly in noisy conditions. With functional magnetic resonance imaging it was found that these linguistic visual cues are sufficient to activate auditory cortex in normal hearing individuals in the absence of auditory speech sounds. Two further experiments suggest that these auditory cortical areas are not engaged when an individual is viewing nonlinguistic facial movements but appear to be activated by silent meaningless speechlike movements (pseudospeech). This supports psycholinguistic evidence that seen speech influences the perception of heard speech at a prelexical stage.     

Exit of the pre-TCR from the ER/cis-Golgi is necessary for signaling differentiation, proliferation, and allelic exclusion in immature thymocytes

A major issue is whether surface expression of the pre-TCR is necessary for signaling the development of immature thymocytes. To address this question, we generated transgenic mice expressing a TCRbeta chain that had a strong endoplasmic reticulum (ER) retrieval signal (TCRbetaER) and that was expressed intracellularly but failed to reach the cell surface. In TCRbetaER transgenic mice, there was a failure of allelic exclusion. Also, the transgene failed to rescue the developmental defects observed in TCRbeta-null mice. In contrast, TCRbeta transgenes with a mutant ER retrieval sequence or lacking this sequence signaled efficient allelic exclusion and suppressed the TCRbeta-/- defect. These data show that exit of the pre-TCR from the ER/cis-Golgi is required for progression through the double-negative thymocyte checkpoint.     

Comparison of calcium phosphate cement mixture and pure calcium hydroxide as direct pulp-capping agents

This review reports the different genetic factors that have been identified either as risk factor for Alzheimer's disease (AD) or directly causing the disease. First are reviewed epidemiological data and biological mechanisms about the apoplipoprotein E gene allele epsilon 4 that is a major risk factor for Alzheimer's disease. The second part describes the mutations responsible for early-onset autosomal dominant AD found in three different genes. The gene located on chromosome 21 encodes the amyloid precusor protein (APP). The presenilin 1 and presenilin 2 genes, located on chromosome 14 and 1 respectively, encode not yet known membrane proteins.     

Carbon monoxide and water vapor contamination of compressed breathing air for firefighters and divers

Compressed breathing air, used in self-contained breathing apparatus (SCBA) by firefighters and other categories of workers as well as by recreational and commercial divers, is prepared with the aid of high-pressure compressors operating in the range of 5000 psig. There have been reports of unexplained deaths of SCUBA divers and anecdotal accounts of decreased time to exhaustion in firefighters using SCBAs. Compressed breathing air has been found to contain elevated levels of carbon monoxide (CO) and water vapor that are consistent with carboxyhemoglobin (COHb) poisoning and freezing of the user's regulator on the breathing apparatus. The Coburn-Forster-Kane equation (CFK equation) was used to estimate COHb levels at rest and at maximum exercise when exposed to different levels of CO in contaminated breathing air. The results demonstrated that, at maximum exercise, the COHb ranged from 6.0 to 17% with the use of 1 to 4 SCBA cylinders contaminated by 250 ppm CO. Standard operating procedures have been developed at the Montreal Fire Department to minimize the risk of compressed breathing air contamination. Results of the quality analysis/quality control program indicate that implementation of these procedures has improved the quality of the compressed breathing air. Recommendations are made for improvement of the air testing procedures mandated by the Canadian CAN3 180.1-M85 Standard on Compressed Breathing Air and Systems.     

The branch site adenosine is recognized differently for the two steps of pre-mRNA splicing

The functional groups responsible for branch site identity in the two steps of pre-mRNA splicing as well as for spliceosome assembly were tested by incorporation of site-specific modifications at the branch site of a pre-mRNA. These results show that recognition of the adenosine occurs early in complex formation and that the branch site adenosine is recognized differently before the first step and for the second step. Further, direct UV cross-linking with these modified RNAs was used to identify a factor whose interaction was dependent upon the identity of the branch site nucleotide.     

The block to HIV-1 envelope glycoprotein-mediated membrane fusion in animal cells expressing human CD4 can be overcome by a human cell component(s)

Membrane fusion mediated by interaction of the human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein with the human CD4 molecule generally requires that the CD4 be expressed on a human cell. The failure of murine or simian cells expressing human CD4 to form syncytia upon mixing with cells expressing envelope glycoprotein could not be corrected by expression of both molecules at extremely high surface levels using vaccinia virus expression vectors. Video fluorescence microscopic analysis of fluorescent dye transfer between fusing cells indicated that the block occurred at the level of membrane fusion between individual pairs of cells. To gain insight into the basis for this fusion block, we tested the ability of fluorescent probe cells expressing envelope glycoprotein to fuse with transient animal x human hybrid giant cells expressing human CD4. The hybrid giant cells were generated either by low-pH-induced fusion of vaccinia-infected cells or by CD4/HIV-1 envelope glycoprotein-mediated cell fusion. We observed that envelope glycoprotein-expressing probe cells efficiently fused with CD4-expressing animal x human hybrid giant cells, independent of whether the CD4 was originally expressed on the animal or on the human cell. Fusion did not occur with CD4-expressing giant cells derived from animal cells alone. These results indicate that the fusion block is not due to dominant inhibitory components in the animal cell. Rather, they suggest that human cells contain an additional component(s) which, when transferred to the CD4-bearing animal cell, confers the ability to undergo membrane fusion mediated by the HIV-1 envelope glycoprotein.