Change search
ReferencesLink to record
Permanent link

Direct link
Essential conserved active site residues in class Ia ribonucleotide reductase from Escherichia coli: identification of radical reaction intermediates and radical transfer intermediates
Stockholm University.
1999 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The enzyme ribonucleotide reductase catalyzes the reduction of ribonucleotides to the corresponding deoxyribonucleotides, the building blocks of DNA. The class Ia enzyme from Escherichia coli consists of two homodimeric subunits denoted protein R1 and protein R2, each inactive alone. Protein R1 contains the active site and allosteric effector sites. Protein R2 contains a stable tyrosyl radical and a dinuclear iron-oxo site essential for catalysis. A radical based reaction mechanism and a radical transfer pathway between R1 and R2 has been proposed.

The three conserved active site residues Glu441, Asn437 and Ser224 were studied. Their functions in catalysis were adressed with site-directed mutagenesis. The mutant proteins were characterized with biochemical, biophysical and structural methods. A carboxylate at position 441 is the minimum requirement for catalysis. Glu441 contributes to substrate binding. Its role as a general base in the first part of the reaction mechanism and as a general acid in the second part of the reaction mechanism is corroborated. The residues Asn437 and Ser224 are essential for catalysis and participate in the first part of the reaction.

E441Q is a suicidal protein converting the normal substrate into a mechanism-based inhibitor. Consecutive reactions with at least three intermediates were identified. The intermediates are free radical species, a cysteine-localized radical, a nucleotide-localized radical and a tyrosyl radical in the radical transfer pathway were identified. This is the first direct evidence for kinetically coupled radical transfer and reaction intermediates in ribonucleotide reductase.

Place, publisher, year, edition, pages
Stockholm: Stockholm University , 1999. , 69 p.
National Category
Biochemistry and Molecular Biology
Research subject
Molecular Biology
URN: urn:nbn:se:su:diva-64235ISBN: 91-7153-975-1OAI: diva2:456378
Public defence
1999-09-17, 13:00
Härtill 4 uppsatser och 2 appendixAvailable from: 2011-11-14 Created: 2011-11-14Bibliographically approved

Open Access in DiVA

No full text

By organisation
Stockholm University
Biochemistry and Molecular Biology

Search outside of DiVA

GoogleGoogle Scholar
The number of downloads is the sum of all downloads of full texts. It may include eg previous versions that are now no longer available

Total: 17 hits
ReferencesLink to record
Permanent link

Direct link