High resolution footprinting of a type I methyltransferase reveals a large structural distortion within the DNA recognition site

The type I DNA methyltransferase M.EcoR124I is a multi-subunit enzyme that binds to the sequence GAAN6RTCG, transferring a methyl group from S-adenosyl methionine to a specific adenine on each DNA strand. We have investigated the protein-DNA interactions in the complex by DNase I and hydroxyl radical footprinting. The DNase I footprint is unusually large: the protein protects the DNA on both strands for at least two complete turns of the helix, indicating that the enzyme completely encloses the DNA in the complex. The higher resolution hydroxyl radical probe shows a smaller, but still extensive, 18 bp footprint encompassing the recognition site. Within this region, however, there is a remarkably hyper-reactive site on each strand. The two sites of enhanced cleavage are co-incident with the two adenines that are the target bases for methylation, showing that the DNA is both accessible and highly distorted at these sites. The hydroxyl radical footprint is unaffected by the presence of the cofactor S-adenosyl methionine, showing that the distorted DNA structure induced by M.EcoR124I is formed during the initial DNA binding reaction and not as a transient intermediate in the reaction pathway.     

Effects of hypertonia on voltage-gated ion currents in freshly isolated hippocampal neurons, and on synaptic currents in neurons in hippocampal slices

A NAD-dependent mannitol dehydrogenase (MtlD) was purified to homogeneity from P. fluorescens DSM50106 and the N-terminal amino acid sequence was determined. An oligonucleotide deduced from this peptide sequence was used as a probe to isolate the mannitol dehydrogenase gene (mtlD) from a genomic library of P. fluorescens. Nucleotide sequence analysis of a 1.8 kb NruI fragment containing the entire mtlD gene revealed an open reading frame of 1482 bp encoding a protein with a calculated molecular weight of 54.49 kDa. The enzyme shared a high similarity with a mannitol dehydrogenase from Rhodobacter sphaeroides and a putative mannitol dehydrogenase of Saccharomyces cerevisae with an overall identity in amino acid sequence of 44% and 42%, respectively, whereas the similarity to mannitol-1-phosphate dehydrogenases of Escherichia coli or Enterococcus faecalis was only about 23% of identical amino acids. By construction of inducible expression plasmids the specific activity of the mannitol dehydrogenase synthesized in E. coli was increased from 0.02 U (mg protein)(-1) to 10 U (mg protein)(-1). After fusion of six histidine codons to the 3' end of mtlD gene and expression in E. coli active mannitol dehydrogenase could be purified in a two-step procedure by affinity chromatography using a Ni2+ matrix column. The purified enzyme exhibited a specific activity of 46 U (mg protein)(-1) and was shown to be a polyol dehydrogenase with a broad substrate spectrum oxidizing efficiently mannitol, sorbitol and arabitol.     

Significance of adrenal hormones in brain energy metabolism

Intensity of glycolysis was studied under conditions of immobilization stress and adrenalectomy as well as with administration of adrenaline and hydrocortisone. The processes of aerobic glycolysis, activity of enzymes of the tricarboxylic acid cycle and mitochondria respiratory chain, content of total, free and protein-bound glycogen, glucose-I-phosphate, glucose-6-phosphate, pyruvate, alpha-ketoglutarate, cytrate and macroergs in the brain are shown to depend on the level of adrenal hormones producing a multilateral effect on a series of energy metabolism links.     

Analysis of the quality of clinical diagnosis from generalized findings of the pathologoanatomic service

A statistical analysis of generalized data of the pathoanatomical service on quality of clinical diagnosis in curative-prophylactic institutions in 54 administrative territories of the RSFSR was carried out. The structure (extensive indices) and frequency (intensive indices)of erroneous clinical diagnoses referring to the most important classes of diseases were identified. As to the structure of indices and frequency of clinico-anatomic disparities the first place was occupied by oncological diseases (20.1+/-0.11 and 14.2+/-0.22%), the second--by infectious diseases (16.5+/-0.1 and 13.0+/-0.34%), the third--by diseases of the digestive system (14.6+/-0.09 and 13.0+/-0.33%), the forth--by diseases of the urogenital system (14.0+/-0.09 and 12.2+/-0.49%), the fifth--by disease of the respiratory system (12.7+/-0.09 and 10.6+/-0.24%), the sixth--by diseases of the cardiovascular system (11.1+/-0.08 and 8.0+/-0.14%). The recommendation is put forward to carry on annually a complex satistical analysis of extensive and intensive indices of erroneous clinical diagnoses demonstrating the quality of clinical diagnosis in therapeutic institutions of a given administrative territory.     

Specific glucocorticoid receptor in the iris--ciliary body of the rabbit

The cytoplasm of the iris--ciliary body of the rabbit contains a receptor capable of specifically binding dexamethasone. This binding protein has a high affinity for dexamethasone (average KD = 2.0 X 10(-8) M), a low capacity (average 4.8 X 10(-13) mol of steroid bound per milligram of protein), and extreme heat sensitivity; it exhibits a pattern of competition virtually identical to that obtained with glucocorticoid receptors from other tissues and shows characteristic physicochemical behavior in various salt concentrations. The demonstration of a specific dexamethasone receptor in the iris--ciliary body provides the first direct biochemical evidence that these tissues may function as a target organ for glucocorticoids.     

Persistent acute renal failure with renal artery stenosis: cure following reconstructive arterial operation

A case of acute renal failure is reported in which renal function returned after surgical correction of renal artery stenosis to the sole functioning kidney. Acute renal failure associated with renal vascular lesions is reviewed. Although often clinically apparent vascular obstruction may be asymptomatic and should be considered in cases of prolonged acute renal failure.     

Factors affecting serum oxytetracycline levels in beef calves

Fifteen Aberdeen Angus steers, 295-364 kg, were dosed with either 4.4 or 11 mg of oxytetracycline hydrochloride/kg im. The antimicrobial activity of the serum was determined periodically, and the resulting data were treated statistically to determine the sources of variation. Variance in serum levels of oxytetracycline activity was attributed to dose, time of bleeding, order of dosing, animal, and assay. The total variance component was proportionately greater for the 11-mg/kg dose than for the 4.4-mg/kg dose. Animal variance increased with the higher dose level of oxytetracycline. The influence of dose on serum level was tested by applying a t test to the mean serum levels and their standard deviations at each bleeding time. The 4.4- and 11-mg/kg serum levels were significantly different (p less than 0.01) at all bleeding times. The 4.4-mg/kg serum levels mutliplied by 2.5 were not significantly different (p less than 0.05) from the 11-mg/kg serum levels at all bleeding times.