Characterization of human phosphoserine aminotransferase involved in the phosphorylated pathway Of L-serine biosynthesis

In the present study, we first report two forms of human phosphoserine aminotransferase (PSAT) cDNA (HsPSATalpha and HsPSATbeta). HsPSATalpha has a predicted open reading frame comprising 324 amino acids, encoding a 35.2 kDa protein (PSATalpha), whereas HsPSATbeta consists of an open reading frame comprising 370 amino acids that encodes a 40 kDa protein (PSATbeta). PSATalpha is identical with PSATbeta, except that it lacks 46 amino acids between Val(290) and Ser(337) of PSATbeta, which is encoded by the entire exon 8 (138 bp). Both PSATalpha and PSATbeta can functionally rescue the deletion mutation of the Saccharomyces cerevisiae counterpart. Reverse transcriptase-PCR analysis revealed that the expression of PSATbeta mRNA was more dominant when compared with PSATalpha mRNA in all human cell lines tested. PSATbeta was easily detected in proportion to the level of mRNA; however, PSATalpha was detected only in K562 and HepG2 cells as a very faint band. The relative enzyme activity of glutathione S-transferase (GST)-PSATbeta expressed in Escherichia coli appeared to be 6.8 times higher than that of GST-PSATalpha. PSAT mRNA was expressed at high levels (approx. 2.2 kb) in the brain, liver, kidney and pancreas, and very weakly expressed in the thymus, prostate, testis and colon. In U937 cells, the levels of PSAT mRNA and protein appeared to be up-regulated to support proliferation. Accumulation of PSAT mRNA reached a maximum in the S-phase of Jurkat T-cells. These results demonstrate that although two isoforms of human PSAT can be produced by alternative splicing, PSATbeta rather than PSATalpha is the physiologically functional enzyme required for the phosphorylated pathway, and indicate that the human PSAT gene is regulated depending on tissue specificity as well as cellular proliferation status with a maximum level expression in the S-phase.