Species differences in the sites of cleavage of pro-lactase to lactase supports lack of selective pressure

The pro-sequences in pro-lactase-phlorizin hydrolase (LPH) are needed for l actase to proceed past the ER, but are irrelevant as to the enzymatic activ ities. Hence, in all species removal of the pro-sequences (or most of them) must take place after the ER. Contrary to this, the details of the removal of these pro-sequences are to be expected to differ in the various species , since they are not subjected to selective pressure. Using site-directed m utagenesis we investigated processing in rabbit. The first cleavage occurs by furin (or furin-like PCs) and takes place at R-A-A-R-349 in the pro-sequ ence, generating the known 180 kDa intermediate. Replacing R-349 by Q resul ts in a mutant which is not cleaved but nevertheless transported to the cel l surface as demonstrated by immunofluorescence. Further processing of eith er the 180 kDa intermediate or the mutant is not directly mediated by furin -like PCs, but involves (also) other proteases. These results demonstrate t hat formation of the 180 kDa intermediate, consistently found only in rabbi ts, but not in man, is not essential for lactase transport: in all likeliho od lack of selective pressure has led to species-specific processing of pro -LPH. (C) 1999 Elsevier Science B.V. All rights reserved.