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Biomedical subjects

A Platz

Publications and source records attributed to A Platz.

85 records · Page 5Linked to original sources
PubMed1984

Mutationally altered ribonucleotide reductase from Escherichia coli: characterization of mutations isolated on multicopy plasmids.

The Escherichia coli ribonucleotide reductase genes (nrd genes) were mutagenized at random. Point mutations were introduced in vitro into a recombinant nrd plasmid. Transformants were initially screened for altered tolerance toward the drug hydroxyurea and further characterized by enzymatic and immunological methods. The screening procedure could pick out defects in either of the two subunits of ribonucleotide reductase. Cells carrying the nrd plasmid pPS2 were earlier shown to have levels of ribonucleotide reductase molecules that were 10 to 20 times higher than those in wild-type cells. We now demonstrate that the enzymatic activity in gently lysed pPS2-containing cells on cellophane disks is six times higher than in wild-type cells. Supplementation of the pPS2-containing lysates with a purified thioredoxin system results in a further 4.5-fold stimulation of the enzymatic activity, which implies a functional shortage of the electron donor system(s) for ribonucleotide reduction in pPS2-containing cells.

Cloning, Molecular↗
PubMed1983

Enzymic modification of a tyrosine residue to a stable free radical in ribonucleotide reductase.

Protein B2, a subunit of ribonucleotide reductase from Escherichia coli, contains in its active form a tyrosyl free radical as part of the polypeptide chain and a dimeric iron center that stabilizes the radical. The enzyme depends on this radical for its catalytic activity. Treatment with hydroxyurea scavenges the radical without disturbing the iron center and, thereby, results in an inactive form of the subunit, B2/HU. A second inactive form, apoB2, lacking both the radical and the iron center, is obtained by treatment of B2 with 8-hydroxyquinoline. Here we describe an enzyme activity in extracts from E. coli that transforms the catalytically inactive B2/HU form into the active B2 subunit by regeneration of the tyrosyl radical. This reaction requires the presence of oxygen, dithiothreitol, and Mg2+ and does not proceed through apoB2. Under anaerobic conditions, we obtained evidence for a second activity in the bacterial extract that destroys the free radical and transforms B2 into B2/HU. We suggest that this novel type of protein modification is functionally related to the synthesis of deoxyribonucleotides and DNA.

Apoproteins↗
PubMed1980

Construction and characterization of hybrid plasmids containing the Escherichia coli nrd region.

Recombinant plasmids containing all or part of the genetic region of Escherichia coli coding for the two subunits of ribonucleoside diphosphate reductase (proteins B1 and B2) were constructed with the aid of the multicopy plasmid pBR322. Two of these plasmids (pPS1 and pPS2) appeared to carry both a regulator and the complete structural information for the enzyme and, after transformation of E. coli, directed a 10- to 20-fold overproduction of both proteins B1 and B2. The other plasmids (pPS101 and pPS201) carried structural information for only protein B2. Cells carrying pPS1 and pPS2 showed a 5- to 500-fold increased resistance against the drug hydroxyurea. This establishes that in E. coli the inhibition of deoxyribonucleic acid synthesis by hydroxyurea is fully explained by its action on ribonucleotide reductase.

DNA, Recombinant↗
PubMed1969

Evoked potentials: modifications by classical conditioning.

Visual evoked potentials to a positive discriminative stimulus change systematically during sensory conditioning and extinction. Changes due to conditioning are manifested in the increased amplitude of the late component of the evoked response. This effect is attenuated during extinction and reappears after reconditioning.

Adult↗
PubMedDate not supplied

Control of severe hemorrhage using C-clamp and pelvic packing in multiply injured patients with pelvic ring disruption.

OBJECTIVES: Evaluation of diagnostic and therapeutic workup in multiply injured patients with pelvic ring disruption and hemorrhagic shock. DESIGN: Prospective study. PATIENTS: Twenty consecutive multiply injured patients (ISS: 41.2 +/- 15.3 points) with pelvic ring disruption and hemorrhagic shock. INTERVENTION: A C-clamp was used for primary stabilization of the pelvic ring instability. In patients with persistent or massive hemorrhage, laparotomy and pelvic packing were performed. Consecutive measurements of blood lactate levels during the early period after injury. MAIN OUTCOME MEASUREMENTS: Lactate, mortality. RESULTS: A C-clamp was applied in all patients within 57.4 +/- 30.6 minutes of arrival. Fourteen patients underwent laparotomy with pelvic packing for control of hemorrhage, three patients additional resuscitation thoracotomy (aortic clamping: n = 2). Four patients died of exsanguinating hemorrhage during the first 5.4 +/- 3.3 hours from arrival, one patient because of septic multi-organ failure twenty-three days after injury (total mortality: 5/20; 25 percent). Lactate levels at admission were elevated in all patients (5.1 +/- 2.6 mmol/l). Increased blood lactate levels (4.8 +/- 1.7 mmol/l) (+71 percent; p < 0.05) were observed in survivors undergoing laparotomy compared with survivors without laparotomy (2.8 +/- 1.1 mmol/l). In contrast, hemoglobin (7.0 +/- 2.6 g/dl versus 7.9 +/- 2.2 g/dl) and hematocrit (21.4 +/- 6.4 percent versus 23.2 +/- 6.8 percent) were similar in both groups. In patients who died during the first hours after admission, lactate levels were elevated (8.6 +/- 2.5 mmol/l) compared with survivors (4.2 +/- 1.8 mmol/l) and increased further. CONCLUSIONS: Sequential measurements of blood lactate levels during the early period after injury may provide a more rapid and reliable estimation of true severity of hemorrhage than routinely used parameters. Pelvic packing in addition to pelvic ring fixation with a C-clamp allows for effective control of severe hemorrhage in multiply injured patients with pelvic ring disruption.

Adolescent↗
PubMedDate not supplied

The Ras farnesylation inhibitor BZA-5B increases the resistance to cisplatin in a human melanoma cell line.

Ras proteins have been implicated in transducing cellular responses to DNA damaging agents. We used BZA-5B, an inhibitor of Ras-farnesylation, to examine the role of Ras in cellular sensitivity to cisplatin. A human melanoma cell line (224) with a Gln61Arg mutation in N-ras was used for these studies. We report that BZA-5B treated cells show an increased resistance to cisplatin. BZA-5B treatment decreased the number of cells showing in situ DNA fragmentation and increased cell viability and clonogenic survival after cisplatin treatment. Further experiments showed that cisplatin induction of the immediate early genes c-jun and p21cip1 was not affected by BZA-5B. Finally, we show that cisplatin causes only weak activation of Jun N-terminal kinase (JNK) in a human melanoma cell line. We conclude that inhibition of Ras function decreases the sensitivity of human melanoma cells to cisplatin-induced cell death.

Alkyl and Aryl Transferases↗