Gene_locus Report for: ratno-Ces1e

The cDNA of the rat carboxylesterase ES-3 encodes a polypeptide with 561 amino acid residues including a cleavable signal peptide at the N-terminus. The processed polypeptide shows over 90% sequence identity to mouse egasyn (ES-22); its calculated pI (5.5) matches the value determined for purified liver ES-3. The product expressed in COS cells migrates in native gels in the region of ES-3 and is similarly active on acetanilide. It is retained in the cells, as predicted from its C-terminus HTEL, and bears a single endo-H sensitive oligosaccharide chain. The nonglycosylated form expressed in the presence of tunicamycin is also intracellular, but substantially less active.

Resident proteins of the endoplasmic reticulum lumen are continuously retrieved from an early Golgi compartment by a receptor-mediated mechanism. The sorting or retention sequence on the endoplasmic reticulum proteins is located at the C-terminus and was initially shown to be the tetrapeptide KDEL in mammalian cells and HDEL in Saccharomyces cerevisiae. The carboxylesterases are a large family of enzymes primarily localized to the lumen of the endoplasmic reticulum. Retention sequences in these proteins have been difficult to identify due to atypical and heterogeneous C-terminal sequences. Utilizing the polymerase chain reaction with degenerate primers, we have identified and characterized the C-termini of four members of the carboxylesterase family from rat liver. Three of the carboxylesterases sequences contained C-terminal sequences (HVEL, HNEL or HTEL) resembling the yeast sorting signal which were reported to be non-functional in mammalian cells. A fourth carboxylesterase contained a distinct C-terminal sequence, TEHT. A full-length esterase cDNA clone, terminating in the sequence HVEL, was isolated and was used to assess the retention capabilities of the various esterase C-terminal sequences. This esterase was retained in COS-1 cells, but was secreted when its C-terminal tetrapeptide, HVEL, was deleted. Addition of C-terminal sequences containing HNEL and HTEL resulted in efficient retention. However, the C-terminal sequence containing TEHT was not a functional retention signal. Both HDEL, the authentic yeast retention signal, and KDEL were efficient retention sequences for the esterase. These studies show that some members of the rat liver carboxylesterase family contain novel C-terminal retention sequences that resemble the yeast signal. At least one member of the family does not contain a C-terminal retention signal and probably represents a secretory form.

We report biochemical, immunological, and genetic studies which demonstrate that an accessory protein with the essential features of mouse egasyn is complexed with and stabilizes a portion of beta-glucuronidase in microsomes of rat liver. The accessory protein exists as a complex with beta-glucuronidase since it coprecipitates with beta-glucuronidase after treatment of extracts with a specific beta-glucuronidase antibody. The two proteins are associated by noncovalent bonds since they are easily dissociated at elevated temperatures. Only 20-25% of total liver accessory protein is complexed with microsomal beta-glucuronidase. The remainder exists as a free form. The molecular weight of the accessory protein is 61 to 63 kDa depending upon the rat strain of origin. This protein, like mouse egasyn, has esterase catalytic activity and is concentrated in microsomes. The accessory protein is genetically polymorphic with at least four alleles. Combined biochemical and genetic evidence indicates it is identical with esterase-3 of the rat. Also, both mouse egasyn and rat esterase-3 react with antisera to egasyn and to rat esterase-3, indicating they are homologous proteins. Several inbred rat strains lack microsomal beta-glucuronidase. The same strains lack the accessory protein, suggesting that stabilization of beta-glucuronidase in rat microsomes requires egasyn.

Egasyn-beta-glucuronidase complex is located at the luminal site of liver microsomal endoplasmic reticulum. When organophosphorus insecticides (OP) are incorporated into the liver microsomes, they become tightly bound to egasyn, a carboxylesterase isozyme, and subsequently, beta-glucuronidase (BG) is dissociated and released into blood. Consequently, the increase in plasma BG activity becomes a good biomarker of OP exposure. Thus, the single administration of EPN (O-ethyl O-p-nitrophenylphenylphosphonothioate), acephate and chlorpyrifos increased plasma BG activity in approximately 100-fold the control level in rats. The increase in plasma BG activity after OP exposure is a much more sensitive biomarker of acute OP exposure than acetylcholinesterase (AChE) inhibition.

The laboratory rat (Rattus norvegicus) is an indispensable tool in experimental medicine and drug development, having made inestimable contributions to human health. We report here the genome sequence of the Brown Norway (BN) rat strain. The sequence represents a high-quality 'draft' covering over 90% of the genome. The BN rat sequence is the third complete mammalian genome to be deciphered, and three-way comparisons with the human and mouse genomes resolve details of mammalian evolution. This first comprehensive analysis includes genes and proteins and their relation to human disease, repeated sequences, comparative genome-wide studies of mammalian orthologous chromosomal regions and rearrangement breakpoints, reconstruction of ancestral karyotypes and the events leading to existing species, rates of variation, and lineage-specific and lineage-independent evolutionary events such as expansion of gene families, orthology relations and protein evolution.

The cDNA of the rat carboxylesterase ES-3 encodes a polypeptide with 561 amino acid residues including a cleavable signal peptide at the N-terminus. The processed polypeptide shows over 90% sequence identity to mouse egasyn (ES-22); its calculated pI (5.5) matches the value determined for purified liver ES-3. The product expressed in COS cells migrates in native gels in the region of ES-3 and is similarly active on acetanilide. It is retained in the cells, as predicted from its C-terminus HTEL, and bears a single endo-H sensitive oligosaccharide chain. The nonglycosylated form expressed in the presence of tunicamycin is also intracellular, but substantially less active.

Resident proteins of the endoplasmic reticulum lumen are continuously retrieved from an early Golgi compartment by a receptor-mediated mechanism. The sorting or retention sequence on the endoplasmic reticulum proteins is located at the C-terminus and was initially shown to be the tetrapeptide KDEL in mammalian cells and HDEL in Saccharomyces cerevisiae. The carboxylesterases are a large family of enzymes primarily localized to the lumen of the endoplasmic reticulum. Retention sequences in these proteins have been difficult to identify due to atypical and heterogeneous C-terminal sequences. Utilizing the polymerase chain reaction with degenerate primers, we have identified and characterized the C-termini of four members of the carboxylesterase family from rat liver. Three of the carboxylesterases sequences contained C-terminal sequences (HVEL, HNEL or HTEL) resembling the yeast sorting signal which were reported to be non-functional in mammalian cells. A fourth carboxylesterase contained a distinct C-terminal sequence, TEHT. A full-length esterase cDNA clone, terminating in the sequence HVEL, was isolated and was used to assess the retention capabilities of the various esterase C-terminal sequences. This esterase was retained in COS-1 cells, but was secreted when its C-terminal tetrapeptide, HVEL, was deleted. Addition of C-terminal sequences containing HNEL and HTEL resulted in efficient retention. However, the C-terminal sequence containing TEHT was not a functional retention signal. Both HDEL, the authentic yeast retention signal, and KDEL were efficient retention sequences for the esterase. These studies show that some members of the rat liver carboxylesterase family contain novel C-terminal retention sequences that resemble the yeast signal. At least one member of the family does not contain a C-terminal retention signal and probably represents a secretory form.

We report biochemical, immunological, and genetic studies which demonstrate that an accessory protein with the essential features of mouse egasyn is complexed with and stabilizes a portion of beta-glucuronidase in microsomes of rat liver. The accessory protein exists as a complex with beta-glucuronidase since it coprecipitates with beta-glucuronidase after treatment of extracts with a specific beta-glucuronidase antibody. The two proteins are associated by noncovalent bonds since they are easily dissociated at elevated temperatures. Only 20-25% of total liver accessory protein is complexed with microsomal beta-glucuronidase. The remainder exists as a free form. The molecular weight of the accessory protein is 61 to 63 kDa depending upon the rat strain of origin. This protein, like mouse egasyn, has esterase catalytic activity and is concentrated in microsomes. The accessory protein is genetically polymorphic with at least four alleles. Combined biochemical and genetic evidence indicates it is identical with esterase-3 of the rat. Also, both mouse egasyn and rat esterase-3 react with antisera to egasyn and to rat esterase-3, indicating they are homologous proteins. Several inbred rat strains lack microsomal beta-glucuronidase. The same strains lack the accessory protein, suggesting that stabilization of beta-glucuronidase in rat microsomes requires egasyn.