NOVEL 39-KDA PHOSPHORIBOSYLPYROPHOSPHATE SYNTHETASE-ASSOCIATED PROTEIN OF RAT-LIVER - CLONING, HIGH SEQUENCE SIMILARITY TO THE CATALYTIC SUBUNITS, AND A NEGATIVE REGULATORY ROLE

The rat liver phosphoribosylpyrophosphate (PRPP) synthetase exists as complex aggregates composed of the 34-kDa catalytic subunits (PRS I an d II) and other 39- and 41-kDa proteins (Kita, K., Otsuki, T., Ishizuk a, T., and Tatibana, M. (1989) J. Biochem. (Tokyo) 105, 736-741), whic h are termed here PRPP synthetase-associated proteins (PAPs). We have cloned the cDNA for the major one of 39 kDa (PAP39) from a rat liver c DNA library. Nucleotide sequencing showed that the clone encoded 356 a mino acids containing sequences of all five peptides derived from PAP3 9. Surprisingly, the deduced amino acid sequence is markedly similar t o those of the 34-kDa catalytic subunits. Excluding two regions (about 45 residues in total), PAP39 has a 48% identity with PRS I. Northern analysis detected a major 1.9-kilobase transcript in all 16 rat tissue s examined, and the relative amounts of PAP39 mRNA to PRS I mRNA varie d with tissues. Covalent cross-linking experiments gave definitive evi dence for molecular interaction of PAP39 with the catalytic subunits. Immunoprecipitation experiments revealed that all the catalytic subuni ts existed as complexes containing PAP39. When PAPs were eliminated fr om the rat liver enzyme complex by gel filtration in the presence of 1 M MgCl2, a lyotrope, or by mild tryptic treatment, the enzyme activit y of the remaining catalytic subunits increased. Based on these result s, we propose that PAP39, the major component of PAPs, plays a negativ e regulatory role in PRPP synthesis and hence is an important factor c ontrolling nucleotide syntheses in general.