PRODUCTION OF THE R2 SUBUNIT OF RIBONUCLEOTIDE REDUCTASE FROM HERPES-SIMPLEX VIRUS WITH PROKARYOTIC AND EUKARYOTIC EXPRESSION SYSTEMS - HIGHER ACTIVITY OF R2 PRODUCED BY EUKARYOTIC CELLS RELATED TO HIGHER IRON-BINDING-CAPACITY

The R2 subunit of ribonucleotide reductase from herpes simplex virus t ype 2 was overproduced with prokaryotic and eukaryotic expression syst ems. The recombinant R2 purified by a two-step procedure exhibited a 3 -fold higher activity when produced in eukaryotic cells. Precise quant ification of the R2 concentration at each step of the purification ind icated that the activity was not altered during the purification proce dure. Moreover, we have observed that the level of R2 expression, in e ukaryotic cells as well. as in prokaryotic cells, did not influence R2 activity. Extensive characterization of the recombinant R2 purified f rom eukaryotic and prokaryotic expression systems has shown that both types of pure R2 preparations were similar in their 76 kDa dimer conte nts (more than 95%) and in their ability to bind the R1 subunit. Howev er, we have found that the higher activity of R2 produced in eukaryoti c cells is more probably related to a higher capability of binding the iron cofactor as well as a 3-fold greater ability to generate the tyr osyl free radical.