Renaturation and stabilization of the telomere-binding activity of Saccharomyces Cdc13(451-693)p by L-arginine

Production of recombinant proteins can be valuable in studying their biolog ical functions. However, recombinant proteins expressed in Escherichia coli sometimes form undesirable insoluble aggregates. Solubilization and renatu ration of these aggregates becomes a problem that one needs to solve. Here we used recombinant Cdc13(451-693)p as example to show the presence of L-ar ginine during renaturation greatly enhanced the renaturation efficiency. Cd c13p is the single-stranded telomere-binding protein of yeast Saccharomyces cerevisiaei The telomere-binding domain has been mapped within amino acids 451-693 of Cdc13p, Cdc13(451-693)p. Recombinant Cdc13(451-693)p was expres sed in E. coli as insoluble protein aggregates. Purification of insoluble C dc13(451-693)p was achieved by denaturing the protein with 6 M guanidine-HC l and followed by Ni-nitrilotriacetic acid agarose column chromatography. R enaturation of Cdc13(451-693)p to the active form was achieved by dialyzing denatured protein in the presence of L-arginine. Moreover, the presence of L-arginine was also helped in maintaining the telomere-binding activity of Cdc13(451-693)p. Taking together, L-arginine might have a general applicat ion in renaturation of insoluble aggregates. (C) 2001 Academic Press.