Isolation of a cDNA encoding the rat liver S‐adenosylmethionine synthetase

Abstract

We have isolated cDNA clones encoding the rat liver S-adenosylmethionine synthetase by means of immunological screening from a phage Agt 11 expression library containing cDNA synthesized from adult rat liver poly(A)-RNA. The amino acid sequence deduced from the cDNA indicates that the rat liver enzyme for this protein contains 397 amino acid residues and has a molecular mass of 43 697 Da. The deduced amino acid sequence of rat liver S-adenosylniethionine synthetase was 68 Yo similar to those of yeast S-adenosylmethionine synthetases encoded by two unlinked genes S A M l and SAM2. The rat liver S-adenosylmethionine synthetase also shows 52%0 similarity with the deduced amino acid sequence of the MetK gene encoding the S-adenosylmethionine synthetase in Escherichia coli.

Analysis of the 5′ non‐coding region of rat liver S‐adenosylmethionine synthetase mRNA and comparison of the Mr deduced from the cDNA sequence and the purified enzyme

“…On the other hand, no fragment was extended when Poly(A)' RNA from kidney was used. A cDNA clone for a kidney AdoMet synthetase has been reported, that in Northern analysis was able to detect two mRNA species in this tissue, but did not hybridize to the liver RNA [13]. Since the liver and kidney AdoMet synthetase cDNA sequences seem to be different, the result obtained with the kidney sample on primer ex,tension was expected.…”

“…Since the liver and kidney AdoMet synthetase cDNA sequences seem to be different, the result obtained with the kidney sample on primer ex,tension was expected. The primer extension analysis also determines in a precise way the ocsurrence of only one mRNA species coding for this protein in rat liver and further excludes the possibility of an alternative transcriptional initiation site, as has been proposed to take place in kidney [13].…”

“…DNA from positive phage was isolated by the plate lysate method [6]. The DNA was digested with EcoRI endonuclease and the inserts subcloned into the EcoRI site of PUC 18, The insert was digested with Haelll, Alul, Sacl, EcoRl and ClaI according to the restriction map already described [5], and that shown in Fig. I Sequencing of both strands was performed by the dideoxy chain termination method [7].…”

“…It has been shown that both forms are constituted by the same polypeptide chain, which has an apparent molecular mass of 48.5 kDa on polyacrylamide gel electrophoresis [3,4]. Recently, Horikawa et al [5] have isolated a 1.8 kb cDNAcoding for rat liver AdoMet synthetase. The deduced amino acid sequence corresponds to a protein with an Mr of 43.7 kDa instead of the 48.5 kDa described for the purified enzyme subunit.…”

DNA methylation and histone acetylation of rat methionine adenosyltransferase 1A and 2A genes is tissue-specific

“…Blots were prehybridized and hybridized as described [20]. Rat MAT1A full length cDNA [21] and a 1147 base pairs cDNA fragment of rat MAT2A [5,22] were labeled with [a-32 P]dCTP (3000 Ci/mmol; Amersham Pharmacia Biotech) by random priming using the Megaprime DNA labeling system (Amersham). The amount and quality of the loaded RNA samples were evaluated by ethidium bromide staining of the gels.…”