Nucleotide sequence of cDNA coding for rat liver pI 6.1 esterase (ES-10), a carboxylesterase located in the lumen of the endoplasmic reticulum

Abstract

A commercial rat liver cDNA library in lambda gt11 was screened with a rabbit antiserum to native pI 6.1 esterase (ES-10). The inserts of the immunoreactive clones were short (0.9-1.1 kbp). One of these was used as a probe to rescreen the library, yielding 30 clones, two of which contained relatively long (approx. 1.9 kbp) and widely overlapping cDNA inserts. They did not contain the first two nucleotide residues of the initiator codon, nor the 5′-end untranslated portion of the mRNA. These were derived from a home-made rat liver cDNA library in lambda gt11, screened with an oligonucleotide corresponding to the 5′-end of the already known cDNA sequence. The nucleotide sequence consists of 48 bp of 5′-end non-coding region, 1695 bp of coding region and 212 bp of 3′-end non-coding region including a 20 bp poly(A) tail. The signal peptide and the mature protein subunit are 18 and 547 residues long respectively. Tyr is confirmed as N-terminal residue. The predicted amino acid sequence is highly similar to those of rabbit liver esterase forms 1 (77% identity) and 2 (56% identity), determined by protein sequencing [Korza & Ozols (1988) J. Biol. Chem. 263, 3486-3495; Ozols (1989) J. Biol. Chem. 264, 12533-12545]. The three enzymes share the Ser and His residues presumed to be part of the active site, four Cys residues and a high proportion of charged side chains at their C-terminus. The C-terminal tetrapeptides of the three esterases (-HVEL, -HIEL and -HTEL for pI 6.1 and forms 1 and 2 esterases respectively) are reminiscent of, but not identical with, the localization signal identified in other proteins of the endoplasmic-reticulum lumen (-KDEL in animal cells [Munro & Pelham (1987) Cell 48, 899-907]; -HDEL in yeast [Pelham, Hardwick & Lewis (1988) EMBO J. 7, 1757-1762]). We still lack direct evidence to decide whether or not these C-terminal tetrapeptides commit esterases to reside in the endoplasmic reticulum. In that case the antepenultimate residue (D, V, I or T) would be only weakly stringent, and some sequences primed by H instead of K would be recognized in animal as well as in yeast cells.