Phosphophoryns (PP), the major noncollagenous proteins (NCPs) in dentin, are believed to play a crucial role in mineral nucleation and hydroxyapatite growth during dentin mineralization. Previously we identified two mature rat PP transcripts, one coding for a 240 amino acid protein (designated as PP(240)) (H.H. Ritchie, L.-H. Wang, J. Biol. Chem. 271 (1996) 21695-21698), and another coding for a 171 amino acid protein (PP(171)) (H. Ritchie, L. Wang, Biochim. Biophys. Acta 1493 (2000) 27-32). We now have identified a third novel dentin sialoprotein (DSP)-PP cDNA transcript that encodes a 523 amino acid protein (PP(523)) with typical PP characteristics including DSS and DS motifs suitable as potential casein kinase I and II phosphorylation sites. Based on amino acid composition, the PP(523) protein product is identical to native rat HP2. We also show that the PP(523) sequence is identical to the corresponding genomic DNA sequence. Taken together, the existence of multiple DSP-PP transcripts, each significantly different from the other in net negative charge, suggests that dentin mineralization processes may be under fine-tune control by these PP protein isoforms.