Association of Bile‐Salt‐Dependent Lipase with Membranes of Human Pancreatic Microsomes

Abstract

Immunolocalization studies indicated that, in contrast to other enzyme markers of human pancreatic secretion, bile-salt-dependent lipase (BSDL) was partly but specifically associated with endoplasmic reticulum membranes. In microsomes, temperature-induced phase separation using Triton X-I 14 elucidated the partition of BSDL between the aqueous phase and the detergent-rich phase containing hydrophilic and membrane proteins, respectively. The size of the membrane-associated BSDL (approx. 100 kDa) is compatible with that of the fully processed enzyme. Fucosylated 0-and N-linked oligosaccharide structures were detected by means of specific lectins. The membrane-associated BSDL might therefore be released from membranes between the trurzs-Golgi compartment (where terminal fucose residues were added) and the zymogen granules where BSDL was mainly found in the soluble fraction. Even though BSDL associated with membranes was enzymically active, it appeared less efficient than the soluble form. The association of BSDL with membranes was pH-dependent and optimal association occurred between pH 5 -6. The membrane-associated BSDL was released by KBr which suggests that the association of BSDL with microsomal membranes involves ionic interactions. Lipid-protein interactions are probably not involved in this association as BSDL did not associate with liver microsome membranes. We attempted to characterize the putative ligand and showed that BSDL and a 94-kDa protein, immunologically related to a glucose-regulated protein of 94 kDa (Grp94), were co-immunoprecipitated by specific antibodies directed against each individual species. It is suggested that the biogenesis of the human pancreatic BSDL involves an association with intracellular membranes and that its folding may be assisted by molecular chaperones.

Endocytosis of Secreted Carboxyl Ester Lipase in a Syndrome of Diabetes and Pancreatic Exocrine Dysfunction

“…The immature, low molecular weight forms of both CEL variants co-distributed with a minor pool of p58 in the fast-sedimenting ER factions (fractions 6 -9). Moreover, they co-distributed with the ER chaperone GRP94 in agreement with the previously reported association of CEL with a GRP94-containing, membrane-bound, multiprotein folding complex (10,32). The somewhat wider distribution profile of CEL-MUT could indicate that this protein was more prone to associate with membranes intracellularly as well as extracellularly ( Fig.…”

“…CEL is kept in close proximity to intracellular membranes during its transport from the endoplasmic reticulum (ER) to post-Golgi compartments in the acinar cells (10). Its membrane association is due to an interaction with a multiprotein complex that contains the chaperone GRP94 (glucose-regulated protein of 94 kDa) (11,12).…”

Control of pancreatic bile-salt-dependent-lipase secretion by the glucose-regulated protein of 94kDa (Grp94)

“…Because genistein inhibits the formation and the release of cargo vesicles in the trans-Golgi network [47] one may suspect that the BSDL dissociates from the membrane folding complex in this latter compartment. This, in part, corroborates the fact that the membrane-associated BSDL was N-and O-glycosylated and appeared to be released from membranes after its glycosylation was completed in the trans-Golgi compartment [3,4]. Altogether, these results suggested that the dissociation of secretioncompetent molecules of BSDL from the folding complex occurs after the trans-Golgi compartment, probably in the transGolgi network, which is already known to segregate vesicle populations in cells [48].…”

“…BSDL molecules that cannot associate with Grp94 are unstable and cannot be secreted. Therefore Grp94 [3,4] may assist the sorting of BSDL towards the secretion pathway. Under these circumstances, the fraction of enzyme that does not associate with Grp94 (or with the folding complex) could be stored within the ER before entering another folding cycle, as described for folding mechanisms involving calnexin [40], and being targeted towards secretion.…”

“…The enzyme is synthesized within the endoplasmic reticulum (ER) of acinar cells, and then follows the secretory pathway of these cells to be secreted as a component of the pancreatic juice [2]. However, in contrast with other pancreatic enzymes, such as α-amylase, colipase-dependent lipase, chymotrypsinogen and pancreatic stone protein, BSDL has been shown to be associated with membranes of the ER and the Golgi apparatus [3,4]. This association involves a membrane multiprotein folding complex, and one of these proteins with which BSDL interacts is the chaperone known as the glucose-regulated protein of 94 kDa (Grp94) [4].…”