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Biomedical subjects

D R Winters

Publications and source records attributed to D R Winters.

5 recordsLinked to original sources
PubMed2000

The simian virus 40 core origin contains two separate sequence modules that support T-antigen double-hexamer assembly.

Using subfragments of the simian virus 40 (SV40) core origin, we demonstrate that two alternative modules exist for the assembly of T-antigen (T-ag) double hexamers. Pentanucleotides 1 and 3 and the early palindrome (EP) constitute one assembly unit, while pentanucleotides 2 and 4 and the AT-rich region constitute a second, relatively weak, assembly unit. Related studies indicate that on the unit made up of pentanucleotide 1 and 3 and the EP assembly unit, the first hexamer forms on pentanucleotide 1 and that owing to additional protein-DNA and protein-protein interactions, the second hexamer is able to form on pentanucleotide 3. Oligomerization on the unit made up of pentanucleotide 2 and 4 and the AT-rich region is initiated by assembly of a hexamer on pentanucleotide 4; subsequent formation of the second hexamer takes place on pentanucleotide 2. Given that oligomerization on the SV40 origin is limited to double-hexamer formation, it is likely that only a single module is used for the initial assembly of T-ag double hexamers. Finally, we discuss the evidence that nucleotide hydrolysis is required for the remodeling events that result in the utilization of the second assembly unit.

Adenosine Triphosphate↗
PubMed2000

Phosphorylation of simian virus 40 T antigen on Thr 124 selectively promotes double-hexamer formation on subfragments of the viral core origin.

Cell cycle-dependent phosphorylation of simian virus 40 (SV40) large tumor antigen (T-ag) on threonine 124 is essential for the initiation of viral DNA replication. A T-ag molecule containing a Thr-->Ala substitution at this position (T124A) was previously shown to bind to the SV40 core origin but to be defective in DNA unwinding and initiation of DNA replication. However, exactly what step in the initiation process is defective as a result of the T124A mutation has not been established. Therefore, to better understand the control of SV40 replication, we have reinvestigated the assembly of T124A molecules on the SV40 origin. Herein it is demonstrated that hexamer formation is unaffected by the phosphorylation state of Thr 124. In contrast, T124A molecules are defective in double-hexamer assembly on subfragments of the core origin containing single assembly units. We also report that T124A molecules are inhibitors of T-ag double hexamer formation. These and related studies indicate that phosphorylation of T-ag on Thr 124 is a necessary step for completing the assembly of functional double hexamers on the SV40 origin. The implications of these studies for the cell cycle control of SV40 DNA replication are discussed.

Amino Acid Substitution↗
PubMed1997

Automated procedure for estimation of blood cholinesterase activities in rabbits.

An automated procedure for estimation of blood cholinesterase activities was evaluated and used to determine baseline cholinesterase activities in rabbits. In addition, the kinetics of cholinesterase inhibition and recovery from diazinon challenge were determined by use of this procedure. The procedure developed is a modification of the colorimetric Ellman technique, adapted for use with the Instrumentation Laboratory Multistat III Plus Centrifugal Analyzer. The substrate used in this technique was acetylthiocholine. Baseline cholinesterase activities in domestic rabbits were plasma cholinesterase, 360 mU/ml; red blood cell cholinesterase, 2,658 mU/ml; and whole blood cholinesterase, 1,332 mU/ml. Rabbits were challenged with 0.3 LD50 (40 mg/kg) of diazinon, and blood samples were obtained at hourly, then daily intervals. Cholinesterase activity in these samples was determined to monitor the effect of the cholinesterase inhibitor diazinon in vivo, as estimated by the Multistat III Plus Centrifugal Analyzer procedure. Plasma cholinesterase activity decreased to 4% within 6 h, whereas red blood cell cholinesterase activity decreased to 51%. By 10 days, plasma and red blood cell cholinesterase activities had returned to 100 and 91%, respectively, of pre-challenge activities.

Animals↗
PubMed1976

use of F(ab')2 fragment in experimental skin transplantation for burns.

In a series of experiments involving approximately 500 animals, burned rats were administered the F(ab')2 fragment of antidonor IgG to study the survival of incompatible skin allografts. No prolongation was noted when treatment consisted of antidonor F(ab')2 fragment alone. This was true whether the fragment was raised against donor spleen cells of donor skin, whether the skin allograft was partial or full thickness and whether the systemic treatment of the recipients was preceded by a period of incubation of the graft in F(ab')2 fragment or not. Significant prolongation of survival was noted when the fragment was raised against both donor and recipient histocompatibility antigen and when the donor skin was incubated in steroids as well as in F(ab')2 fragment prior to grafting. Additional findings included the confirmation of prolonged allograft survival by increasing burn size and the prolongation of allograft survival by the presence of intact eschar.

Animals↗