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An ultrafiltration assay for nucleotide binding to ribonucleotide reductase
- Source :
- Analytical biochemistry. 189(1)
- Publication Year :
- 1990
-
Abstract
- Direct partition through ultrafiltration was applied to develop a method for the study of nucleotide binding to ribonucleotide reductase from Escherichia coli. The assay involved a 0.5- to 1-min centrifugation step where bound and unbound nucleotides are separated over an ultrafiltration membrane. No effects were seen due to hyperconcentration of protein at the membrane surface. The method was verified by measuring binding of dATP, ATP, dTTP, dGTP, and GDP at 25 and 4 degrees C with dissociation constants ranging from 0.1 to 80 microM. The results were in good agreement with earlier data obtained by other techniques and extend our knowledge in the case of ATP and dGTP binding at 25 degrees C.
- Subjects :
- Biophysics
Ultrafiltration
Centrifugation
Plasma protein binding
Biology
medicine.disease_cause
Biochemistry
Adenosine Triphosphate
Deoxyadenine Nucleotides
Bacterial Proteins
Ribonucleotide Reductases
medicine
Escherichia coli
Thymine Nucleotides
heterocyclic compounds
Nucleotide
Molecular Biology
chemistry.chemical_classification
Nucleotides
DGTP binding
Deoxyguanine Nucleotides
Proteins
Cell Biology
Dissociation constant
Kinetics
Membrane
Ribonucleotide reductase
chemistry
Guanosine Triphosphate
Protein Binding
Subjects
Details
- ISSN :
- 00032697
- Volume :
- 189
- Issue :
- 1
- Database :
- OpenAIRE
- Journal :
- Analytical biochemistry
- Accession number :
- edsair.doi.dedup.....b7ffb5d78b80e84b4fa61a6a9568bd82