HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via CrossRef Citing Articles via Google Scholar Google Scholar Articles by Conner, J. Articles by Marsden, H. Search for Related Content PubMed PubMed Citation Articles by Conner, J. Articles by Marsden, H. Agricola Articles by Conner, J. Articles by Marsden, H.

Identification of structural domains within the large subunit of herpes simplex virus ribonucleotide reductase

MRC Virology Unit, Church Street, Glasgow G11 5JR, U.K.

The large subunit (R1) of herpes simplex virus (HSV) ribonucleotide reductase is a bifunctional protein consisting of a unique N-terminal protein kinase domain and a ribonucleotide reductase domain. Previous studies showed that the two functional domains are linked by a protease sensitive site. Here we provide evidence for two subdomains, of 30K and 53K, within the reductase domain. The two fragments, which were produced by limited proteolysis and were resistant to further degradation, remained tightly associated in a complex containing two molecules of each. They were capable of binding the R2 subunit of HSV ribonucleotide reductase with approximately the same affinity as the intact protein but the complex did not complement the small subunit (R2) to give an active enzyme. At low concentrations (0.4 µg/ml) of trypsin or V8 protease, cleavage between the subdomains was prevented by the presence of the N-terminal protein kinase domain. At higher protease concentrations (1 µg/ml) the N-terminal domain is extensively proteolysed and the 30K and 53K domains were generated. Identical results were obtained using purified R1 isolated from infected cell extracts or following expression in Escherichia coli. The origin of the two domains was investigated by N-terminal sequencing of the 53K fragment and by examining their reactivity with a panel of R1-specific monoclonal antibodies which we isolated and epitope mapped for that purpose. The trypsin cleavage site was found to lie between arginine 575 and asparagine 576, and proteolysis in this region was not prevented by the presence of R2 or the nonapeptide YAGAVVNDL. We propose that the ribonucleotide reductase region of HSV R1 exists in a two domain structure, and that the interdomain linking region is protected by the unique N terminus.

Received 6 June 1994; accepted 17 August 1994.

This article has been cited by other articles:

				J Conner The unique N terminus of herpes simplex virus type 1 ribonucleotide reductase large subunit is phosphorylated by casein kinase 2, which may have a homologue in Escherichia coli J. Gen. Virol., June 1, 1999; 80(6): 1471 - 1476. [Abstract]

				Y. Langelier, L. Champoux, M. Hamel, C. Guilbault, N. Lamarche, P. Gaudreau, and B. Massie The R1 Subunit of Herpes Simplex Virus Ribonucleotide Reductase Is a Good Substrate for Host Cell Protein Kinases but Is Not Itself a Protein Kinase J. Biol. Chem., January 16, 1998; 273(3): 1435 - 1443. [Abstract] [Full Text] [PDF]