PubMed HealthSearch

Biomedical subjects

M Paloheimo

Publications and source records attributed to M Paloheimo.

8 recordsLinked to original sources

Production of cyclomaltodextrin glucanotransferase of Bacillus circulans var. alkalophilus ATCC21783 in B. subtilis.

The cyclomaltodextrin glucanotransferase (CGTase, E.C. 2.4.1.19) gene from an alkalophilic Bacillus circulans var. alkalophilus ATCC21783 was cloned into Escherichia coli and B. subtilis. When cloned from E. coli to B. subtilis, the entire insert containing the CGTase gene was, depending on the plasmid construction, either unstable or the recombinant B. subtilis did not secrete the enzyme in significant amounts. To achieve efficient enzyme production in B. subtilis, the gene was placed under the control of the B. amyloliquefaciens alpha-amylase promoter. In one of the constructions, both the promoter and the signal sequence of the gene were replaced with those of B. amyloliquefaciens, whereas in another construction only the promoter area was exchanged. The recombinant B. subtilis clones transformed with these plasmid constructions secreted CGTase into the culture medium 14 times as much as did the parental strain in shake flask cultures. In fermentor cultures in an industrially feasible medium the enzyme production was substantially higher, yielding 1.2 g/l of CGTase, which is about 33 times the amount of the enzyme produced by the parental strain in corresponding fermentations. Both of the plasmid constructions were stable when grown over 50 generations without antibiotic selection.

Amino Acid Sequence

Monoclonal antibodies against core and cellulose-binding domains of Trichoderma reesei cellobiohydrolases I and II and endoglucanase I.

Cellulases from Trichoderma reesei form an enzyme group with a common structural organization. Each cellulase enzyme is composed of two functional domains, the core region containing the active site and the cellulose-binding domain (CBD). To facilitate the specific detection of each domain, monoclonal antibodies (mAb) against cellobiohydrolase I (CBHI), cellobiohydrolase II (CBHII) and endoglucanase I (EGI) were produced. Five mAb were obtained against CBHI, ten against CBHII and eight against EGI. The location of the antigenic epitope for each antibody was mapped by allowing the antibodies to react with truncated cellulases, synthesized from deleted cDNA in Saccharomyces cerevisiae. Proteolytic fragments of Trichoderma cellulases, obtained by papain digestion, were used to confirm the results. Specific antibodies were detected against the core and the CBD epitopes for all three cellulases. Using the truncated enzymes, it was possible to locate the epitopes to a reasonably short region within the protein. To obtain a quantitative assay for each enzyme, a specific mAb against each antigen was chosen, based on the affinity to the corresponding antigen on Western-blot staining and on filter blots of the cellulolytic yeasts. The mAb were used to quantitative the corresponding enzymes in T. reesei culture medium. Specific quantitation of each cellulase enzyme has not been possible by biochemical assays or using polyclonal antibodies, due to their cross-reactions. Now, these mAb can be specifically used to recognize and quantitate different domains of these three important cellulolytic enzymes.

Antibodies, Monoclonal

The conserved terminal region of Trichoderma reesei cellulases forms a strong antigenic epitope for polyclonal antibodies.

The specificity of polyclonal antibodies (Pab) raised against Trichoderma reesei cellulases has been studied. cDNAs lacking regions coding for certain functional domains were produced by preparing series of 3'-end deletions from the cDNAs for two cellobiohydrolases, CBH I and CBH II, and an endoglucanase, EG I. The proteins coded by the full length cDNAs and the truncated proteins coded by the deleted cDNAs were expressed in yeast Saccharomyces cerevisiae, under the control of the ADC1 promoter. Each polyclonal antiserum showed cross-reactivity with other cellulases. Pabs for CBH I and CBH II both recognized EG I. Pab for EG I strongly recognized both CBH I and CBH II. By analyzing the truncated proteins, we found that these antibodies were almost entirely directed against the conserved tail of the cellulase enzymes.

Antibodies, Fungal

Periocular anaesthesia: technique, effectiveness and complications with special reference to postoperative ptosis.

The effectiveness of periocular anaesthesia and its complications were examined in 100 successive cataract operations. The patients were divided into 3 groups according to the duration of ocular compression with an Autopressor device after administration of periocular anaesthesia. In the control group, no compression was used (C-O, n = 36 patients). In the other two groups, compression was used for 10 (C-10, n = 32) and for 20 (C-20, n = 32) min. No differences in globe or orbicular akinesia were found between the groups. At 10 min, immobilisation of the globe in different directions was attained in 60.1-84.5% of the patients. Compression for an additional 10 min did not significantly improve the akinesia. In contrast, the hitherto undescribed loss of light perception increased with time: 15 patients at 10 min and 22 at 20 min were unable to see light. Chemosis and haematomas in the upper eyelid occurred more often in C-0 than in the other 2 groups. One day postoperatively the average palpebral aperture was smaller in C-0 than in the other two groups. The frequent postoperative ptosis (74.3% on the 1st day) decreased rapidly, but on postoperative day 7, 9 patients still had ptosis. In only one patient was ptosis still recognizable at 6 weeks postoperatively. No serious complications occurred. This study demonstrates that periocular anaesthesia with ocular compression is a suitable method for cataract surgery.

Adult

Quantitative surface electromyography (qEMG): applications in anaesthesiology and critical care.

During general anaesthesia and in lowered vigilance states such as after major trauma and during heavy sedation or analgesic medication, patients' ability to communicate with their surroundings is limited. Subjective intuitional interpretation may be the only means to ascertain a patient's emotional state, mood, and pain perception. Electromyographic detection and quantification of minimal and covert facial mimic muscle activity in anaesthesiology and critical care was an interesting concept worth further evaluation. In this study, the behaviour of quantitative surface-detected electromyographic activity (qEMG) was investigated during common anaesthetic events, post-operatively, and in volunteers as well as in experimental animals. A review of the methodology includes the necessary details for reproduction of the studies, including computerized processing of numerical data available in the commercial equipment. Results from the monitoring of 218 patients, seven volunteers and 31 rats are discussed. Conclusions are based on 32 testable null-hypotheses, the earlier documented literature and the author's own experience. The qEMG signal was derived from two electrodes placed on the frontal area and on the mastoid process behind the ipsilateral ear. After amplification, the signal was filtered to obtain a portion containing electrical activity between 60-300 Hz, which was considered to represent electromyographic activity. The signals were thereafter full-wave rectified and averaged with a 1-s time constant. The output of the processing unit consisted of a graphics display and a numeric computer output. A variety of clinical conditions and drug effects were studied in order to evaluate the method's applicability in research and in routine anaesthetic practice. The facial muscles turned out to be less sensitive to the effects of neuromuscular blocking drugs than the hand muscles, the normal monitoring site of neuromuscular transmission. Although muscle relaxants had a suppressing effect on spontaneous EMG activity, they did not abolish the ability of facial muscles to react to noxious stimuli. Also abdominal muscles retained this capability, though presumably through a different motor mechanism. Depolarizing and non-depolarizing neuromuscular blocking drugs had disparate effects on spontaneous EMG activity; succinylcholine seemed to facilitate the qEMG amplitude during recovery of the block. Quantitative surface electromyography revealed impending arousals during lightening anaesthesia, although this was not a consistent phenomenon. Inadequate anaesthesia was always reflected by an increase in facial qEMG, albeit this often was also evident to the naked eye. The arousal at the end of anaesthesia was always associated with an abrupt increase in facial qEMG activity, which often was preceded by a more gradual, predictive rise. Auditory stimulation was also effective in increasing qEMG.(ABSTRACT TRUNCATED AT 400 WORDS)

Anesthesia

Open scavenging systems.

A mathematical model was developed to cover the dynamic characteristics of open scavenging systems. The performance of such a device depends on the expiratory tidal volume (VT), the scavenging flow (Vs) and the mode of exhalational volume build-up. A widely used open scavenging system was tested by using carbon dioxide in oxygen as a marker gas. Capnography revealed gross turbulence in the expiratory reservoir, which tends to increase the polluted gas volumes and loads extra requirements onto the scavenging equipment. Increases in the scavenging flow or the expiratory reservoir volume are the aids in obtaining satisfactory safety margins for pollution-free scavenging of surplus anaesthetic gases. The theoretical basis of a new method to determine the expiratory time constant of the respiratory system is also presented.

Air Pollutants