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Biomedical subjects

J Metzger

Publications and source records attributed to J Metzger.

At least 19 recordsLinked to original sources
PubMed1996

Protein engineering of lantibiotics.

Whereas protein engineering of enzymes and structural proteins nowadays is an established research tool for studying structure-function relationships of polypeptides and for improving their properties, the engineering of posttranslationally modified peptides, such as the lantibiotics, is just coming of age. The engineering of lantibiotics is less straightforward than that of unmodified proteins, since expression systems should be developed not only for the structural genes but also for the genes encoding the biosynthetic enzymes, immunity protein and regulatory proteins. Moreover, correct posttranslational modification of specific residues could in many cases be a prerequisite for production and secretion of the active lantibiotic, which limits the number of successful mutations one can apply. This paper describes the development of expression systems for the structural lantibiotic genes for nisin A, nisin Z, gallidermin, epidermin and Pep5, and gives examples of recently produced site-directed mutants of these lantibiotics. Characterization of the mutants yielded valuable information on biosynthetic requirements for production. Moreover, regions in the lantibiotics were identified that are of crucial importance for antimicrobial activity. Eventually, this knowledge will lead to the rational design of lantibiotics optimally suited for fighting specific undesirable microorganisms. The mutants are of additional value for studies directed towards the elucidation of the mode of action of lantibiotics.

Amino Acid Sequence↗
PubMed1996

Evaluation of household dust mite exposure and levels of specific IgE and IgG antibodies in asthmatic patients enrolled in a trial of immunotherapy.

BACKGROUND: Monitoring the response to immunotherapy entails understanding exposure to relevant allergens. For the major indoor allergens, this requires sampling of dust from the patient's house. The objectives of this study were to measure indoor allergen levels during a controlled trial of dust mite immunotherapy for asthma and to relate these results to serum antibody levels. METHODS: Eighty-eight asthmatic patients with mite allergy from seven geographic areas in the United States were enrolled in and completed a course of immunotherapy with Dermatophagoides extract or placebo control. Sensitization was evaluated by quantitative measurements of IgG and IgE antibodies. Dust samples were assayed for group I mite (Der p 1 and Der f 1), cat (Fel d 1), and cockroach (Bla g 1) allergens by monoclonal antibody-based ELISA. RESULTS: Over the 4 years of the study, each of the houses had at least one sample that contained more than 2 micrograms of group I mite allergen per gram of dust. Mean mite allergen levels, however, varied over a wide range, from 0.2 microgram/gm or less to more than 50 micrograms/gm. IgE antibodies to mite were present in sera from 78% of the patients, whereas IgE antibodies to cat and cockroach allergens were found in sera from 34% and 11% of patients, respectively. Sixty-four percent of the patients had exposure and sensitization to mite, whereas the comparable figure for each of the other allergens was 5%. CONCLUSIONS: Examination of the results suggested that allergen exposure, relative to a trial of immunotherapy, could be expressed as (1) the maximum level found in the house, (2) the percentage of sites having greater than 2 micrograms/gm, or (3) the mean value at the site with the maximum level. This report provides a background for evaluating the clinical results of immunotherapy in these patients and a model for the way in which sensitization and exposure should be monitored in studies of this kind.

Adolescent↗
PubMed1995

Active and inactive transplacement of the M26 recombination hotspot in Schizosaccharomyces pombe.

The ade6-M26 mutation of the fission yeast Schizosaccharomyces pombe creates a meiotic recombination hotspot that elevates ade6 intragenic recombination approximately 10-15-fold. A heptanucleotide sequence including the M26 point mutation is required but not sufficient for hotspot activity. We studied the effects of plasmid and chromosomal context on M26 hotspot activity. The M26 hotspot was inactive on a multicopy plasmid containing M26 embedded within 3.0 or 5.9 kb of ade6 DNA. Random S. pombe genomic fragments totaling approximately 7 Mb did not activate the M26 hotspot on a plasmid. M26 hotspot activity was maintained when 3.0-, 4.4-, and 5.9-kb ade6-M26 DNA fragments, with various amounts of non-S. pombe plasmid DNA, were integrated at the ura4 chromosomal locus, but only in certain configurations relative to the ura4 gene and the cointegrated plasmid DNA. Several integrations created new M26-independent recombination hotspots. In all cases the non-ade6 DNA was located > 1 kb from the M26 site, and in some cases > 2 kb. Because the chromosomal context effect was transmitted over large distances, and did not appear to be mediated by a single discrete DNA sequence element, we infer that the local chromatin structure has a pronounced effect on M26 hotspot activity.

Chromosomes, Fungal↗
PubMed1995

Isolation and characterization of genetically engineered gallidermin and epidermin analogs.

Gallidermin (Gdm) and epidermin (Epi) are highly homologous tetracyclic polypeptide antibiotics that are ribosomally synthesized by a Staphylococcus gallinarum strain and a Staphylococcus epidermidis strain, respectively. These antibiotics are secreted into media and are distinguished by the presence of the unusual amino acids lanthionine, 3-methyllanthionine, didehydrobutyrine, and S-(2-aminovinyl)-D-cysteine, which are formed by posttranslational modification. To study the substrate specificities of the modifying enzymes and to obtain variants that exhibit altered or new biological activities, we changed certain amino acids by performing site-specific mutagenesis with the Gdm and Epi structural genes (gdmA and epiA, respectively). S. epidermidis Tü3298/EMS6, an epiA mutant of the Epi-producing strain, was used as the expression host. This mutant synthesized Epi, Gdm, or analogs of these antibiotics when the appropriate genes were introduced on a plasmid. No Epi or Gdm analogs were isolated from the supernatant when (i) hydroxyamino acids involved in thioether amino acid formation were replaced by nonhydroxyamino acids (S3N and S19A); (ii) C residues involved in thioether bridging were deleted (delta C21, C22 and delta C22); or (iii) a ring amino acid was replaced by an amino acid having a completely different character (G10E and Y20G). A strong decrease in production was observed when S residues involved in thioether amino acid formation were replaced by T residues (S16T and S19T). A number of conservative changes at positions 6, 12, and 14 on the Gdm backbone were tolerated and led to analogs that had altered biological properties, such as enhanced antimicrobial activity (L6V) or a remarkable resistance to proteolytic degradation (A12L and Dhb14P). The T14S substitution led to simultaneous production of two Gdm species formed by incomplete posttranslational modification (dehydration) of the S-14 residue. The fully modified Dhb14Dha analog exhibited antimicrobial activity similar to that of Gdm, whereas the Dhb14S analog was less active. Both peptides were more sensitive to tryptic cleavage than Gdm was.

Amino Acid Sequence↗
PubMed1995

[Pseudotumor calcinosis in renal failure].

Tumoral Calcinosis is a rare clinical entity characterized by the presence of calcified, periarticular masses, as seen on radiographs. A severe case in a patient with chronic renal failure is presented and the complex etiology of this disease is discussed.

Adult↗
PubMed1995

[Small bowel injuries in blunt abdominal trauma--a diagnostic problem!].

AIMS: Injuries of the small bowel following a blunt abdominal trauma are rare (incidence 6%). The diagnosis is difficult and therefore often missed. The aim of this paper is to acquire an useful diagnostic sequence of our own results in comparison to the literature. METHODS: The diagnostic procedures and the time of the final diagnosis of 13 patients with this rare injury have been retrospectively analysed. RESULTS: Two of these 13 patients died in the emergency room. Five of the remaining 11 patients had signs of peritonitis at the first examination. These were confirmed by findings either in peritoneal lavage (3), ultrasound (1) or X-ray (1). Patients without signs of peritonitis revealed no pathological signs at the first examination with these diagnostic tools. Correct diagnosis was achieved on 7/11 patients within 24 hours. The diagnosis of small bowel injury was confirmed by the rest of the patients (n = 4) after an average of 11 days after trauma. DISCUSSION/CONCLUSION: Initial assessment with diagnostic tools as ultrasound, peritoneal lavage or X-ray is poor. Therefore repetitive controls (Nowadays by ultrasound) are indicated. The CT is a valuable tool to detect an organ lesion in hemodynamic stable patients with positive proof of liquid in the ultrasound. Today the value of the diagnostic laparoscopy is not established. It should be considered that a local ischemia due to the blunt trauma can cause a delayed perforation.

Abdomen↗
PubMed1994

Elucidation of the structure of SA-FF22, a lanthionine-containing antibacterial peptide produced by Streptococcus pyogenes strain FF22.

The antibacterial peptide SA-FF22, produced by the pathogen Streptococcus pyogenes strain FF22 was purified and features of its primary and secondary structure were characterised. Mass spectrometry demonstrated the pure peptide had a mass of 2794Da while, amino acid analysis revealed the presence of the unusual, thioether amino acids lanthionine and 3-methyllanthionine; thus SA-FF22 is a member of the group of antibacterial polypeptides termed lantibiotics. Furthermore, amino acid sequencing showed a unique sequence which was blocked at position 23 by a residue of the unsaturated amino acid 2,3-didehydrobutyrine. Carboxypeptidase-Y digestion could be used to demonstrate that serine occupies the C-terminal position only after complete oxidation of the thioether amino acid bridges, suggesting that the three-dimensional structure of the native peptide may prevent access of the enzyme to the C-terminus. Fragmentation of the native peptide with a variety of proteolytic enzymes failed to yield a peptide containing less than all three of the cross-linked lanthionine and methyllanthionine residues and demonstrated that all three thioether bridges overlapped. Analysis of the circular dichroism of SA-FF22 in various concentrations of 2,2,2-trifluoroethanol in water, SDS micelles and in the presence of artificial phospholipid vesicles suggested that there is significant change in its secondary structure from aqueous to lipophilic environments.

Alanine↗
PubMed1994

Factors influencing changes in the signal-averaged electrocardiogram within the first year after a first myocardial infarction.

One hundred twenty-nine patients were prospectively studied after a first myocardial infarction. A first signal-averaged electrocardiogram (SAECG-1) was performed in the acute phase (within 48 hours after onset of symptoms) and a second one (SAECG-2) in the late phase (6 to 18 months after hospital discharge). We studied the influence of nine parameters on the evolution of the signal-averaged electrocardiogram: age, gender, myocardial infarction location, number of diseased coronary vessels, infarct-related coronary artery patency, use of thrombolytic therapy or percutaneous transluminal coronary angioplasty in the acute phase, left ventricular ejection fraction, and recurrence of ischemic events. No follow-up data were available in 15 patients. Of the remaining 114 patients, an ischemic event occurred in 25 (22%). The signal-averaged electrocardiogram remained unchanged in 97 (85%) (remaining normal in 78 and abnormal in 19). It became abnormal in 13 (11.5%) and became normal in 4 (3.5%). In patients with a normal SAECG-1, two factors were associated with the change to an abnormal SAECG-2: (1) an ischemic event occurred in 11 (85%) of 13 patients whose SAECG-2 was abnormal compared with only 13 (17%) of 78 patients whose SAECG-2 remained normal (p < 0.0001), and (2) 100% of patients with an abnormal SAECG-2 had an inferior myocardial infarction compared with 54% of patients with a normal SAECG-2 (p = 0.004).(ABSTRACT TRUNCATED AT 250 WORDS)

Aged↗
PubMed1994

Lipopeptide-polyoxyethylene conjugates as mitogens and adjuvants.

Two lipopeptide analogues of the Escherichia coli lipoprotein rendered water-soluble by polyoxyethylene were tested for mitogenicity in vitro in murine and human B lymphocytes and for adjuvant activity in vivo in mice. These highly amphiphilic lipopeptides retained the biological activity other lipopeptides usually exerted which supports the hypothesis of specific interactions of lipopeptides with membranes of reactive cells. The activation of human B lymphocytes by these lipopeptides was much less pronounced compared to that of murine cells. However, given in combination with anti-CD40 antibodies plus interleukin-4, human B lymphocytes could synergistically be stimulated to proliferate. As an adjuvant, the polyoxyethylene linked lipopeptides were almost as potent as Freund's adjuvants and other basic lipopeptides. Being water-soluble, these novel analogues are easy to apply and they are suitable for field studies as adjuvants when sonication can not usually be provided.

Adjuvants, Immunologic↗
PubMed1994

[1-year follow-up of laparoscopic cholecystectomy in an unselected patient sample. Objective findings and subjective status].

We report on the 1-year follow-up after laparoscopic cholecystectomy in a consecutive series of 116 unselected patients. So far no such follow-up data have been published. The follow-up rate was 94%. All patients have been personally assessed on the base of a standardized follow-up form either at the hospital (58%) or by the general practitioner (42%). Using an adapted modified Visick classification we have found an excellent or good result in 96% of the patients. Although there were some minor dyspeptic complaints and a few abnormal laboratory findings no patient had a suspicion of residual or recurrent gall stones. There has been no reoperation. In conclusion laparoscopic cholecystectomy yields very satisfactory short-term results 1 year after operation even in an unselected population of patients.

Adult↗
PubMed1994

[Is the intravenous cholangiogram of value in the preoperative period?].

The need for a routine preoperative intravenous cholangiogram (IVC) has been controversially discussed. We decided to assess if preoperative criteria such as history, clinical examination or laboratory findings could be used for selective indication for preoperative IVC. In a series of 146 patients with a preoperative IVC before undergoing laparoscopic cholecystectomy, history, clinical findings and laboratory results (bilirubin, transaminases, alkaline phosphatase, amylase) have been correlated with the radiological findings. ERCP was taken as the standard to assess the value of IVC. A normal IVC was quite reliable in excluding any pathology of the bile ducts or common bile duct stones. Thus specificity reached 96% and the negative predictive value was 97%. On the other hand a pathological IVC proved not to be a valid predictor of true pathological alterations. Sensitivity was only 60% and the positive predictive value just 55%. Over all accuracy was quite satisfactory (94%). We could not find a correlation between history, clinical or laboratory findings and the final result as assessed by ERCP. Therefore we could not find any useful parameters to define a selective policy for indication of preoperative IVC. On the other hand the IVC still proved useful to exclude relevant pathological findings. At the time being there is no strong argument for abandoning routine preoperative IVC.

Cholangiography↗
PubMed1993

The 12-lead electrocardiogram in midseptal, anteroseptal, posteroseptal and right free wall accessory pathways.

The 12-lead electrocardiograms of 50 patients with 1 anterogradely conducting accessory pathway were analyzed to obtain characteristics of electrocardiographic findings in the midseptal, anteroseptal, true posteroseptal and right free wall accessory pathway locations. Locations were confirmed by surgery (33 patients) or radiofrequency catheter ablation (17 patients). This study analyzed (1) QRS in the frontal plane, (2) delta wave axis in the frontal plane, (3) the angle between QRS and delta wave axes, (4) the R/S ratio in lead III, (5) negativity of delta wave in inferior leads, and (6) the R/S ratio in precordial leads.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1993

Improvement in left ventricular function by ablation of atrioventricular nodal conduction in selected patients with lone atrial fibrillation.

Left ventricular (LV) function was studied in 30 patients with lone atrial fibrillation (AF) (paroxysmal [n = 27] and persistent [n = 3]) before and after ablation of atrioventricular conduction. In all patients, drug treatment did not control ventricular rate during AF or prevent recurrences of the arrhythmia, or both. LV ejection fraction, and LV end-systolic and end-diastolic, and left atrial dimensions were measured by echocardiography before (mean 7 +/- 10 months, range < 1 to 37) and after (14 +/- 20 months, < 1 to 77) ablation. Before ablation, LV ejection fraction was < or = 50% in 12 patients (group I) and > 50% in 18 (group II). After ablation, LV ejection fraction increased significantly in group I from 43 +/- 8% to 54 +/- 7% (p < 0.0001). There were also significant decreases in LV-end systolic and end-diastolic, and left atrial dimensions. No changes in these parameters were observed in group II. Groups I and II had a significant difference in the duration of AF (group I: mean 11 years, range 8 to 28; and group II: 5 years, 2 to 14) (p < 0.05). No difference was present in age, sex, New York Heart Association functional class for dyspnea, or type of ablation procedure. Thus, some patients with lone AF may show deterioration of LV function, which appears to be related to the duration of the arrhythmia; in these cases, LV function may improve significantly after ventricular rate control is accomplished by ablation of atrioventricular conduction.

Adult↗
PubMed1993

Effects on the signal-averaged electrocardiogram of opening the coronary artery by thrombolytic therapy or percutaneous transluminal coronary angioplasty during acute myocardial infarction.

One hundred twenty-nine patients were retrospectively analyzed and divided into 3 groups according to (1) the presence of a patent artery obtained either spontaneously or after thrombolytic therapy but without percutaneous transluminal coronary angioplasty (PTCA) (group I, n = 83), (2) the presence of a patent artery after opening by PTCA (group II, n = 29), or (3) absence of reperfusion despite thrombolytic therapy or PTCA (group III, n = 17). Thrombolytic therapy was given within 4 hours after onset of symptoms (mean 2.5 +/- 1.0 hours) and PTCA was performed within 24 hours after the onset of symptoms (mean 6 +/- 6 hours). Signal averaging was performed within 24 hours after cardiac catheterization. An abnormal signal-averaged electrocardiogram was present in 10 of 83 (12%) group I, 9 of 29 (31%) group II and 7 of 17 (41%) group III patients (p < 0.05 group I vs II, p < 0.01 group I vs III, no statistical difference group II vs III). Therefore, in contrast to reperfusion by thrombolytic therapy the incidence of abnormalities on the signal-averaged electrocardiogram early after myocardial infarction is not reduced by an early opening of the culprit vessel by PTCA.

Angioplasty, Balloon, Coronary↗
PubMed1993

Echocardiographic wall motion abnormalities and the signal averaged electrocardiogram in the acute phase of a first myocardial infarction.

We studied the relationship between wall motion abnormalities determined by echocardiography and the signal-averaged electrocardiogram in 82 consecutive patients during the acute phase of a first myocardial infarction. An abnormal signal-averaged electrocardiogram was defined as the presence of two of the following criteria: a QRS duration > or = 114 ms, a root mean square voltage (RMS) of the last 40 ms < or = 25 microV and an amplitude signal lower than 40 microV lasting > or = 39 ms. The left ventricle was divided into 13 segments and the contraction pattern divided into akinesia alone (including dyskinesia) (group A), hypokinesia alone (group B) and both hypokinesia and akinesia (group C). An abnormal signal-averaged electrocardiogram was found in 14/82 patients (17%) and was correlated with the persistence of occlusion of the infarct-related vessel (32% vs 9%, P < 0.02). In patients with a patent vessel, the incidence of an abnormal signal-averaged electrocardiogram was 14% in group A, 9% in group B and 0% in group C (NS). In patients with an occluded vessel an abnormal signal-averaged electrocardiogram was found in 10% of group A patients, in 36% in group B patients and in 75% of group C patients (P = 0.05). Our study suggests that the presence of hypokinetic areas during the acute phase of a first myocardial infarction and an abnormal signal-averaged electrocardiogram indicate an occluded infarct-related vessel.

Aged↗
PubMed1993

Infusion of stroma-free cross-linked hemoglobin during acute gram-negative bacteremia.

Twelve dogs were divided into two groups of six each, and were infused with bis-3,5-dibromosalicyl fumarate stroma-free hemoglobin (DBBF-Hb) or albumin. Their responses to an intravenous bolus of Escherichia coli were followed for 4 hr. Bacterial clearance from the blood stream was studied using standard colony counting methodology as well as blood counts, blood chemistries, and clotting factor analysis. There was a significant difference in mean arterial pressure (MAP) over time between DBBF-Hb-treated dogs and those treated with albumin (P < 0.02). While the DBBF-treated dogs had a higher MAP during the 10 min of bacteremia, after 1 hr, there were no longer any appreciable differences between septic dogs treated with DBBF-Hb vs. albumin. Consumption of clotting and natural anticoagulant factors was observed to be similar in both groups, as were endotoxin levels. Blood urea nitrogen (BUN) increased slightly in both groups, while white blood cell counts and clotting factor levels fell in both groups in a similar fashion. There was a more pronounced fall (P < 0.04) in platelet counts in the animals treated with DBBF-Hb. In the dogs treated with DBBF-Hb, there was also a late rise in pCO2 (P < 0.01), a more pronounced fall in pO2, and greater acidosis, which suggested that ventilation perfusion abnormalities may have been exacerbated by DBBF-Hb treatment. Since the exacerbation of respiratory abnormalities was not related to diminished bacterial or endotoxin clearance, the possibility is raised that DBBF-Hb interferes with compensatory respiratory changes during sepsis.

Acute Disease↗