Topology of Binding Sites for Carbamyl Phosphate in Aspartate Transcarbamylase from Escherichia coli
- 1 May 1975
- journal article
- Published by Wiley in European Journal of Biochemistry
- Vol. 54 (1), 293-299
- https://doi.org/10.1111/j.1432-1033.1975.tb04139.x
Abstract
Pyridoxal phosphate, a competitive inhibitor of aspartate transcarbamylase, binds to six sites in the catalytic and to twelve sites in the regulatory subunits of this hexameric protein. The properties of its association to the active sites of the enzyme are very similar to those observed with one of its substrates, carbamyl phosphate. It tightly binds to one half of the sites in the absence of succinate, an analogue of the second substrate. Since pyridoxal phosphate can be linked covalently to the protein by reduction, the distribution of the high affinity binding sites on catalytic trimers was studied after dissociation of modified holoenzyme. Electrophoresis of isolated subunits under non-denaturing conditions revealed four distinct bands, corresponding to trimers containing 0 to 3 pyridoxal phosphate derivatives. The distribution among the four species as a function of ligand concentration in the absence of succinate indicates that in the native oligomer, pyridoxal phosphate (and by extrapolation, carbamyl phosphate) binds to both catalytic trimers, rather than to three sites on a single subunit.Keywords
This publication has 15 references indexed in Scilit:
- Modification of Escherichia coli membranes in the prereplicative phase of phage T4 infection. Specificity of association and quantitation of bound phage proteins.Journal of Biological Chemistry, 1975
- Analysis of bacteriophage T7 early RNAs and proteins on slab gelsJournal of Molecular Biology, 1973
- [41] Techniques to assay repressorsPublished by Elsevier ,1971
- Binding of cytidine triphosphate to aspartate transcarbamylaseBiochemical and Biophysical Research Communications, 1970
- A Discussion of the Regulatory Properties of Aspartate Transcarbamylase from Escherichia coliCurrent Topics in Cellular Regulation, 1970
- The Reliability of Molecular Weight Determinations by Dodecyl Sulfate-Polyacrylamide Gel ElectrophoresisJournal of Biological Chemistry, 1969
- The Purification of Aspartate Transcarbamylase of Escherichia coli and Separation of Its Protein SubunitsJournal of Biological Chemistry, 1967
- DISC ELECTROPHORESIS – II METHOD AND APPLICATION TO HUMAN SERUM PROTEINS*Annals of the New York Academy of Sciences, 1964
- [142] Assay and preparation of pyridoxal phosphateMethods in Enzymology, 1957
- THE ATTRACTIONS OF PROTEINS FOR SMALL MOLECULES AND IONSAnnals of the New York Academy of Sciences, 1949