MUTATIONAL ANALYSIS OF HUMAN LIPOPROTEIN-LIPASE BY CARBOXY-TERMINAL TRUNCATION

We have previously reported a Trp382 (TGG) --> Stop (TGA) mutation tha t causes familial lipoprotein lipase (LPL) deficiency. Expression stud y of the Trp382 --> stop mutant showed that the truncated LPL was cata lytically inactive with a marked reduction in the expressed mass. To i nvestigate the minimal amino-terminal sequence of human LPL required f or an active enzyme, a series of carboxy-terminal (C-terminal) truncat ed LPLs were expressed in vitro, and the enzyme activity, mass, and LP L mRNA levels were analyzed. The lipolytic activity showed a stepwise reduction between LPL-437 (Cys438 --> stop; 68% of normal LPL activity in medium) and LPL-434 (Phe435 --> stop; 3%). Without affecting LPL m RNA levels, LPL mass was reduced with the mutants not larger than LPL- 437. In terms of specific activity, a significant difference was obser ved between LPL-436 (LyS437 --> stop; 88% of that of normal LPL in med ium) and LPL-435 (Val436 --> stop; 22%), implying the importance of th e role of Val436 in LPL action. Furthermore, our results unexpectedly showed that LPL-446 (Ser447 --> stop), which is considered to be a com mon polymorphic form of LPL, exhibited higher activity than normal LPL (185% in medium). These results demonstrate that the C-terminal regio n of human LPL is closely associated with the expression of enzyme mas s and catalytic activity.