Dimerization of lactase-phlorizin hydrolase occurs in the endoplasmic reticulum, involves the putative membrane spanning domain and is required for an efficient transport of the enzyme to the cell surface
Naim, H.Y.; Naim, H.
European Journal of Cell Biology 70(3): 198-208
1996
ISSN/ISBN: 0171-9335 PMID: 8832204 Document Number: 465690
Analysis of the quaternary structure of human intestinal lactase-phlorizin hydrolase (LPH) by chemical cross-linking and sucrose-gradient centrifugation reveals that the brush border form of LPH (LPH-beta; 160-kDa) is a homodimeric molecule. Dimerization ensures in the ER when LPH is still exclusively found as an uncleaved mannoserich precursor (pro-LPH-h; 215-kDa). This is supported by the following observations. (i) Biosynthetically labeled intestinal biopsy specimens as well as transfected COS-1 cells expressing pro-LPH contain monomeric and dimeric forms of pro-LPH-h; the complex glycosylated pro-LPH (pro-LPH-c; 230-kDa) as well as the cleaved mature LPH-beta species in intestinal biopsy samples are discerned exclusively as dimers. (ii) Dimeric forms of pro-LPH-h could be also detected when cells were biosynthetically labeled at 15 degree C, at which temperature the egress of pro-LPH from the ER is blocked. Dimerization is essential for the transport competence of pro-LPH and is strongly associated with the presence of an intact transmembrane domain. Mutant pro-LPH-mact lacking the complete transmembrane domain persists as a monomeric, mannose-rich and transport-incompetent molecule that is not secreted into the exterior milieu, accumulates most likely in the ER and is ultimately degraded. Further, deletion of the cytoplasmic tail in the pro-LPH-ct mutant leads to marked reduction in the proportion of dimeric as well as complex glycosylated pro-LPH-ct. Finally, dimerization is linked to the acquisition of LPH to its biological function, since only dimers of wild type pro-LPH or pro-LPH-ct are enzymatically active, while their monomeric counterparts as well as pro-LPH-mact are not.