Two Plasmodium falciparum ribonucleotide reductase small subunits, PfR2 and PfR4, interact with each other and are components of the in vivo enzyme complex.
Ribonucleotide reductase (RNR) is a tetrameric enzyme, composed of two large (R1) and two small (R2) subunits, which regulates the nucleotide balance in cells by controlling the rate-limiting step for deoxyribonucleotide synthesis. We have identified a second copy of the small subunit gene, termed PfR4, encoding a 324 amino acid residue polypeptide that shares only 25% identity with the previously identified PfR2 small subunit of Plasmodium falciparum. PfR4 expression is cell-cycle-regulated, and the profile of transcript and protein expression corresponds to that of PfR2. A 1.3 kb PfR4 5'-flanking fragment contained a functional promoter activity. We have detected interaction between PfR2 and PfR4 by co-immunoprecipitation experiments. Indirect immunofluorescence analysis showed distinct localization of two small RNR subunits with some colocalization. The association of PfR1 large subunit with PfR4 was detected by GST pull-down assay. This interaction is reduced significantly when using a PfR4 truncated at the COOH terminus, suggesting the involvement of COOH-terminal residues in PfR4-PfR1 interaction. All three RNR subunits co-eluted on a Superose 12 size-exclusion column corresponding to fractions with a molecular mass of around 250 kDa. This suggests the existence of all three RNR subunits in Plasmodium in a native complex of alpha2betabeta' configuration.
Bracchi-Ricard
Valerie
V
Department of Molecular Biology & Microbiology, University of Central Florida, Orlando, FL 32826, USA.
Moe
David
D
Chakrabarti
Debopam
D
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D019143:Evolution, Molecular; D005786:Gene Expression Regulation; D047468:Immunoprecipitation; D008969:Molecular Sequence Data; D046912:Multiprotein Complexes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010963:Plasmodium falciparum / Q000201:enzymology*; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D021381:Protein Transport; D012264:Ribonucleotide Reductases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D014158:Transcription, Genetic / Q000235:genetics
D046912:Multiprotein Complexes; D021122:Protein Subunits; D012264:Ribonucleotide Reductases