PROTEIN DOMAINS IMPLICATED IN INTRACELLULAR-TRANSPORT AND SORTING OF LACTASE-PHLORHIZIN HYDROLASE

The roles of various domains of intestinal lactase-phlorizin hydrolase (pro-LPH) on its folding, dimerization, and polarized sorting are inv estigated in deletion mutants of the ectodomain fused or not fused wit h the membrane-anchoring and cytoplasmic domains (MACT), Deletion of 2 36 amino acids immediately upstream of MACT has no effect on the foldi ng, dimerization, transport competence, or polarized sorting of the mu tant LPH1646MACT, By contrast, LPH1646, an anchorless counterpart of L PH1646MACT, is not transported beyond the ER and persists as a mannose -rich monomer during its entire Life cycle. The further deletion of 87 amino acids generates a correctly folded but transport-incompetent mo nomeric LPH1559MACT mutant. The results strongly suggest that dimeriza tion and transport of pro-LPH implicate a stretch of 87 amino acids in the ectodomain between LPH1646MACT and LPH1559MACT. In addition, dime rization of pro-LPH requires at least two further criteria: (i) a corr ectly folded ectodomain of pro-LPH and (ii) the presence of the transm embrane region. Neither of these requirements alone is sufficient for dimerization. Finally, the sorting of pro-LPH appears to be mediated b y signals located between the cleavage site of pro-LPH and the LPH1646 MACT mutant.