Properties of the deoxyribonucleic acid-binding site of Escherichia coli ribonucleic acid polymerase [proceedings].
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Biomedical subjects
Publications and source records attributed to A D Malcolm.
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The imido ester, methyl acetimidate, which specifically amidinates lysine residues, modifies RNA polymerase core enzyme, leading to rapid loss of activity. Calf thymus DNA partially protects the enzyme against this inactivation, an effect which disappears at high salt concentration. DNA protects 17 +/- 6 lysines from amidination at low salt concentration. The dependence of amidination on methyl acetimidate concentration is examined in the presence of DNA at high and low salt concentration. Analysis of the data suggests a class of approximately 12 lysines which are protected by DNA, consistent with the above estimate. These lysines are approximately 5--10-fold more reactive than most other available lysine residues.
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Left ventricular endomyocardial biopsy specimens were obtained at cardiac catheterisation in 11 patients with mitral-leaflet prolapse, chest pain, and normal coronary arteries. Specimens were examined for birefringence before and after contraction induced by adenosine triphosphate and were also subjected to histochemical analysis. Biopsy specimens from a control group of patients with rigorously defined normal left ventricular function and another control group with poor left ventricular function were also studied. Biopsy specimens from patients with mitral-leaflet prolapse showed a markedly subnormal birefringence response to adenosine triphosphate, with the group mean value intermediate between values for controls with normal and poor left ventricular function. Histochemical studies in the prolapse group also revealed an abnormal pattern, particularly for monoamine-oxidase activity. This is further evidence of a cardiomyopathy in this symptomatic subgroup of patients with mitral-leaflet prolapse.
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1 The treatment of pain of cardiac origin requires a knowledge of the haemodynamic action of the analgesic agents used. 2 The haemodynamic effects of morphine, diamorphine, pavaveretum, pethidine and pentazocine are reviewed. 3 Clinical experience with the new antagonist analgesic buprenorphine is reported. 4 These studies indicate that buprenorphine may be the agent of choice for the relief of severe pain in patients with unstable circulation.
Right atrial hypertension and pericardial effusion developed 2 years after mastectomy for fibrosarcoma. Clinical and echocardiographic features suggested right ventricular tumour. At catheterization the right ventricular cavity was almost obliterated, with an infundibular gradient of 13 mmHg, and biopsy of the mass was attempted. Post-mortem revealed extensive infiltrating and intracavitary right ventricular fibrosarcoma without extracardiac tumour.
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The effect of buprenorphine on the cardiovascular system was examined in 11 patients during the period of reduced cardiac reserve after open-heart surgery. Within 10 minutes of giving the full analgesic dose (5 microgram/kg) intravenously the mean heart rate had fallen significantly by six beats/min. Although in two patients the mean arterial pressure fell by 24 mm Hg, there was no overall change in mean arterial pressure, cardiac output, or peripheral resistance. In a further six patients buprenorphine was used successfully as the sole analgesic after open-heart surgery. Buprenorphine appears to be safer than morphine for use in patients with reduced cardiac reserve and is of similar analgesic efficacy.
Phosphoenolpyruvate carboxylase from the Crassulacean plant Bryophyllum fedtschenkoi has been purified to homogenetity by DEAE-cellulose treatment, (NH4)2SO4 fractionation,, and chromatography on DEAE-cellulose and hydroxyapatite. Poly(ethylene glycol) is required in the extraction medium to obtain maximum enzyme activity. The purified enzyme has a specific activity of about 26 units/mg of protein at 25 degrees C. It gives a single band on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, corresponding to a mol.wt. of 105,000, and gives a single band on non-denaturing gel electrophoresis at pH8.4. Cross-linking studies at pH8.0 indicate that the subunit structure is tetrameric but that the dimer may also be an important unit of polymerization. Gel filtration results at pH6.7 confirm that the native enzyme is tetrameric with a concentration-dependent dissociation to a dimer. The kinetic behaviour is characterized by (i) relatively small variations in maximum velocity between pH5.5 and 9.0 with a double optimum, (ii) a reversible temperature-dependent inactivation between 30 and 45 degrees C, (iii) inhibition by malate, which is pH-sensitive, and (iv) almost Michaelis-Menten behaviour with phosphoenolpyruvate as the varied ligand but sigmoidal behaviour under suitable conditions with malate as the varied ligand. The findings are related to other studies to the possible role phosphoenolpyruvate carboxylase in controlling a circadian rhythm of CO2 fixation.
1. Periodate oxidation of the ribose ring was used to synthesize derivatives of nucleoside triphosphates. 2. These oxidized nucleoside triphosphates. 2. These oxidized nucleoside triphosphates are competitive inhibitors of RNA polymerase. 3. On incubation, together with NaBH4, these oxidized labelled nucleotides are covalently bound to Escherichia coli RNA polymerase. 4. Nucleoside triphosphate substrates decrease the extent of labelling. 5. A lysine residue in an alpha-subunit is labelled. 6. The significance of these results in relation to the location of the nucleotide-binding site is discussed.
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The response to electrocardiographically monitored submaximal exercise stress testing has been studied in 44 patients with mitral leaflet prolapse (MLP). With exercise, ventricular premature contractions occurred in 7, ventricular tachycardia in 1, and atrial fibrillation in 1. Exercise was terminated short of target heart rate in 18 patients, because of chest pain (5), fatigue (7), ventricular arrhythmia (4), dizziness (1) or ST segment depression (1). 23 patients developed postexercise ST segment abnormalities, of whom 5 had 'ischemic' patterns and arteriographically proven coronary artery disease (CAD); among the 18 others, the ST segments were depressed and minimally downsloping in 2, slowly ascending from depressed J point in 3, horizontal for greater than or equal to 80 msec with J depression of less than 1 mm in 12, and cupped in 1. The incidence of arrhythmias provoked by submaximal exercise stress testing in patients with MLP was lower than suggested in previous reports. In all 5 cases where MLP and CAD coexisted, the classical 'ischemic' electrocardiographic response to exercise was not obscured. Even in the absence of CAD, postexercise ST segment abnormalities were common with MLP (18/39 = 46%) and differed from the progressively resolving ST segment deviation characteristic of CAD with angina. Exercise testing can safely be recommended, subject to standard contraindications, in patients with MLP and yields useful information.